Apoptosis In Lung Epithelial Cells Research Articles

D-dopachrome tautomerase contributes to lung epithelial repair via atypical chemokine receptor 3-dependent Akt signaling.

BackgroundEmphysematous COPD is characterized by aberrant alveolar repair. Macrophage migration inhibitory factor (MIF) contributes to alveolar repair, but for its structural and functional homolog D-dopachrome tautomerase (DDT) this is unknown. MIF mediates its effects through CD74 and/or C-X-C chemokine receptors 2 (CXCR2), 4(CXCR4), and possibly 7 (ACKR3). DDT can also signal through CD74, but interactions with other receptors have not been described yet. We therefore aimed at investigating if and how DDT contributes to epithelial repair in COPD.MethodsWe studied effects of recombinant DDT on cell proliferation and survival by clonogenic assay and annexin V-PI staining respectively. DDT-induced signaling was investigated by Western blot. Effects on epithelial growth and differentiation was studied using lung organoid cultures with primary murine or human epithelial cells and incubating with DDT or an ACKR3-blocking nanobody. DDT-ACKR3 interactions were identified by ELISA and co-immunoprecipitation.FindingsWe found that DDT promoted proliferation of and prevented staurosporine-induced apoptosis in A549 lung epithelial cells. Importantly, DDT also stimulated growth of primary alveolar epithelial cells as DDT treatment resulted in significantly more and larger murine and human alveolar organoids compared to untreated controls. The anti-apoptotic effect of DDT and DDT-induced organoid growth were inhibited in the presence of an ACKR3-blocking nanobody. Furthermore, ELISA assay and co-immunoprecipitation suggested DDT complexes with ACKR3. DDT could activate the PI3K-Akt pathway and this activation was enhanced in ACKR3-overexpressing cells.InterpretationIn conclusion, DDT contributes to alveolar epithelial repair via ACKR3 and may thus augment lung epithelial repair in COPD.

Plant-derived exosomal microRNAs inhibit lung inflammation induced by exosomes SARS-CoV-2 Nsp12

Lung inflammation is a hallmark of coronavirus disease 2019 (COVID-19). In this study, we show that mice develop inflamed lung tissue after being administered exosomes released from the lung epithelial cells exposed to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Nsp12 and Nsp13 (exosomesNsp12Nsp13). Mechanistically, we show that exosomesNsp12Nsp13 are taken up by lung macrophages, leading to activation of nuclear factor κB (NF-κB) and the subsequent induction of an array of inflammatory cytokines. Induction of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β from exosomesNsp12Nsp13-activated lung macrophages contributes to inducing apoptosis in lung epithelial cells. Induction of exosomesNsp12Nsp13-mediated lung inflammation was abolished with ginger exosome-like nanoparticle (GELN) microRNA (miRNA aly-miR396a-5p. The role of GELNs in inhibition of the SARS-CoV-2-induced cytopathic effect (CPE) was further demonstrated via GELN aly-miR396a-5p- and rlcv-miR-rL1-28-3p-mediated inhibition of expression of Nsp12 and spike genes, respectively. Taken together, our results reveal exosomesNsp12Nsp13 as potentially important contributors to the development of lung inflammation, and GELNs are a potential therapeutic agent to treat COVID-19.

Avermectin induced DNA damage to the apoptosis and autophagy in human lung epithelial A549 cells

Avermectin (AVM), as a biological insecticide, is widely used in agriculture and forestry production globally. However, inhalation of AVM may pose a risk, and the lung is the direct target, but the cytotoxicity of AVM on human lung cells is still unclear. Here, we attempted to elucidate the cytotoxic effect and molecular mechanism of AVM on human lung A549 cells. The results indicated that AVM inhibits cell proliferation, and enhances programmed cell death (apoptosis and autophagy). In addition, we found the AVM-treated cells showed an obvious drop in mitochondrial membrane potential and LC3-I/II, increased ROS production, DNA double-strand breaks, caspase-3/9 activated, PARP cleaved, cytochrome c and Bax/Bcl-2 content rise. The results showed that AVM induced mitochondria-related apoptosis and autophagy in lung A549 cells. These results indicate that AVM can pose a potential threat to human health by inducing DNA damage and programmed cell death.

Senescence associated long non-coding RNA 1 regulates cigarette smoke-induced senescence of type II alveolar epithelial cells through sirtuin-1 signaling

ObjectiveThe primary aim of our study was to explore the mechanisms through which long non-coding RNA (lncRNA)-mediated sirtuin-1 (SIRT1) signaling regulates type II alveolar epithelial cell (AECII) senescence induced by a cigarette smoke-media suspension (CSM).MethodsPharmacological SIRT1 activation was induced using SRT2104 and senescence-associated lncRNA 1 (SAL-RNA1) was overexpressed. The expression of SIRT1, FOXO3a, p53, p21, MMP-9, and TIMP-1 in different groups was detected by qRT-PCR and Western blotting; the activity of SA-β gal was detected by staining; the binding of SIRT1 to FOXO3a and p53 gene transcription promoters was detected by Chip.ResultsWe found that CSM increased AECII senescence, while SAL-RNA1 overexpression and SIRT1 activation significantly decreased levels of AECII senescence induced by CSM. Using chromatin immunoprecipitation, we found that SIRT1 bound differentially to transcriptional complexes on the FOXO3a and p53 promoters.ConclusionOur results suggested that lncRNA-SAL1-mediated SIRT1 signaling reduces senescence of AECIIs induced by CSM. These findings suggest a new therapeutic target to limit the irreversible apoptosis of lung epithelial cells in COPD patients.

Effect of water-soluble PM10 on the production of TNF-α by human monocytes and induction of apoptosis in A549 human lung epithelial cells.

Long-term exposure to airborne particles of 10 µm and less in size (PM10) in dust can lead to increased risk of diseases such as respiratory, cardiovascular, lung cancer and atherosclerosis. The aim of the study was to evaluate the effects of water-soluble PM10 particles in the Middle East Dust (MED) storm in Ahvaz, Iran, on the production of TNF-α by human monocytes. In addition, we assessed the level of induction of apoptosis in isolated A549 human pulmonary epithelial cells. For this purpose, isolated human blood monocytes (250,000 to 300,000 cell/ ml) as well as isolated human pulmonary A549 epithelial cells (100,0000 cell/ ml) were exposed to various concentrations (62.5, 125, 250, 500µg/ml) of water-soluble PM10 particles for different incubation periods (12, 24, 48h). The results showed that exposure to PM10 particles increased the production of TNF-α in human monocytes and promoted apoptosis induction in A549 cells, in both concentration and incubation of period-dependent manner. In conclusion, airborne dust particles in Ahvaz city contain active compounds capable of increasing production of the pro-inflammatory cytokine, TNF-α, and inducing apoptosis of lung epithelial cells.

Induction of Apoptosis in Human Lung Epithelial Cell by Sphingomonas sp. Shah, a Recently Identified Cell Culture Contaminant.

Sphingomonas sp. Shah is a bacterium that was first isolated from mammalian cell cultures. According to ribotyping data it is very much homologous to the clinically important pathogen Sphingomonas paucimobilis, which has generated pseudo-outbreaks. Using a tissue culture system, Sphingomonas sp. Shah was discovered to induce apoptosis in human lung epithelial carcinoma. Apoptosis of infected cells was determined by numerous criteria including (1) visual alterations in cellular morphology, (2) initiation of nuclear marginalization and chromatin compaction condensation, (3) the attendance of a high percentage of cells with subG1 DNA content, and (4) caspase-3 activation. In the current study we demonstrate the induction of apoptosis in mammalian lung epithelial cells upon infection with Sphingomonas sp. Shah and provide insight into the molecular processes triggering apoptosis.

Hyperoxia induces endoplasmic reticulum stress‑associated apoptosis via the IRE1α pathway in rats with bronchopulmonary dysplasia.

Bronchopulmonary dysplasia (BPD) is the most common chronic lung disease in premature infants, and alveolar dysplasia and pulmonary vascular development disorders are the predominant pathological features. Apoptosis of lung epithelial cells is a key factor in the pathological process of alveolar developmental arrest. Endoplasmic reticulum stress (ERS)-associated apoptosis is a noncanonical apoptotic pathway involved in the development of several pulmonary diseases. Previous studies have demonstrated that protein kinase RNA-like endoplasmic reticulum kinase, inositol-requiring enzyme 1α (IRE1α) and activating transcription factor 6 can initiate the apoptosis signaling pathway mediated by ERS and induce apoptosis of injured cells. Among them, the IRE1α pathway is the most conservative pathway in the unfolded protein response, which serves an important role in a number of pathological environments, to the extent of determining cell fate; however, it is rarely reported in BPD. Based on the establishment of a rat BPD model, the present study verified the activation of ERS in BPD and further confirmed that prolonged ERS inhibited the protective pathway, IRE1α/X-box binding proteins, and activated the proapoptotic pathway, IRE1α/c-Jun N-terminal kinase, to induce the apoptosis of lung epitheliums.

Long non-coding RNA Hsp4 alleviates lipopolysaccharide-induced apoptosis of lung epithelial cells via miRNA-466m-3p/DNAjb6 axis

Acute lung injury (ALI), as a life-threatening syndrome, is mainly characterized with diffuse alveolar injury, excessive pulmonary inflammation, edema and apoptosis of lung epithelial cells. This study investigated the effects of LncRNA Hsp4 (Hsp4, ENSMUST00000175718) on lipopolysaccharide (LPS)-induced apoptosis of MLE-12 cells. In our research, we found that LPS treatment remarkably induced apoptosis of MLE-12 cells and decreased the expression of Hsp4. Overexpression of Hsp4 significantly reversed LPS-induced cell apoptosis through inhibiting mTOR signaling, while suppression of Hsp4 presented opposite effects. Further results showed that Hsp4 positively regulated the expression of miR-466m-3p. Knockdown of miR-466m-3p reversed LPS-induced cell apoptosis via increasing the levels of DNAjb6 which was confirmed to be the target gene of miR-466m-3p. This finding will be helpful for further understanding the critical roles of Hsp4 in ALI and may provide potential targets for ALI diagnosis and treatment.

14-Deoxy-11,12-didehydroandrographolide inhibits apoptosis in influenza A(H5N1) virus-infected human lung epithelial cells via the caspase-9-dependent intrinsic apoptotic pathway which contributes to its antiviral activity

Influenza A virus (IAV) infection represents a global health challenge. Excavating antiviral active components from traditional Chinese medicine (TCM) is a promising anti-IAV strategy. Our previous studies have demonstrated that 14-deoxy-11,12-didehydroandrographolide (DAP), a major ingredient of a TCM herb called Andrographis paniculata, shows anti-IAV activity that is mainly effective against A/chicken/Hubei/327/2004 (H5N1), A/duck/Hubei/XN/2007 (H5N1), and A/PR/8/34 (H1N1) in vitro and in vivo. However, the underlying anti-IAV molecular mechanism of DAP needs further investigation. In the present work, we found that DAP can significantly inhibit the apoptosis of human lung epithelial (A549) cells infected with A/chicken/Hubei/327/2004 (H5N1). After DAP treatment, the protein expression levels of cleaved PARP, cleaved caspase-3, and cleaved caspase-9, and the activities of caspase-3 and caspase-9 in H5N1-infected A549 cells were all obviously downregulated. However, DAP had no inhibitory effect on caspase-8 activity and cleaved caspase-8 production. Meanwhile, the efficacy of DAP in reducing the apoptotic cells was lost after using the inhibitor of caspase-3 or caspase-9 but remained intact after the caspase-8 inhibitor treatment. Moreover, DAP efficiently attenuated the dissipation of mitochondrial membrane potential, suppressed cytochrome c release from the mitochondria to the cytosol, and decreased the protein expression ratio of Bax/Bcl-2 in the mitochondrial fraction. Furthermore, the silencing of caspase-9 reduced the yield of nucleoprotein (NP) and disabled the inhibitory ability of DAP in NP production in A549 cells. Overall results suggest that DAP exerts its antiviral effects by inhibiting H5N1-induced apoptosis on the caspase-9-dependent intrinsic/mitochondrial pathway, which may be one of the anti-H5N1 mechanisms of DAP.

Long Noncoding RNA DLIC Reduces Inflammation, Oxidative Stress and Apoptosis in Lung Epithelial Cells Induced by Lipopolysaccharide (LPS)

The effective improvement of acute lung injury (ALI) plays an important role in reducing the mortality of sepsis. LncRNA was expressed abnormally in inflammatory diseases and restoration of lncRNA could improve the inflammatory injury. However, the protective effects of DLIC on ALI, as well as the regulatory mechanism of DLIC in ALI are still unknown. LPS-induced lung epithelial cells were constructed in vitro to simulate the ALI model. The expression of DLIC in lung epithelial cells and DLIC transfection effects were analyzed by RT-qPCR analysis. The cell viability and cell apoptosis were respectively detected by CCK-8 assay and flow cytometry analysis. The expression of inflammatory factors and oxidative stress factors was determined by the commercial assay kits. The expression of apoptosis-related proteins and TLR4/NF-κB pathway-related proteins was analyzed by western blot analysis. As a result, DLIC expression was decreased in LPS-induced BEAS-2B cells. DLIC overexpression improved the cell viability and inhibited the apoptosis of LPS-induced BEAS-2B cells with the changes of protein expression. DLIC overexpression alleviated the inflammation and oxidative stress by depressing the levels of inflammatory factors and oxidative stress factors. Furthermore, DLIC overexpression inhibited TLR4/NF-κ B pathway. In conclusion, DLIC reduces inflammation, oxidative stress and apoptosis in LPS-induced BEAS-2B cells by suppressing TLR4/NF-κB pathway.

Let-7b-5p is involved in the response of endoplasmic reticulum stress in acute pulmonary embolism through upregulating the expression of stress-associated endoplasmic reticulum protein 1.

The endogenous non-coding microRNA (miRNA) let-7b-5p is highly expressed in the blood of patients with acute pulmonary embolism (PE). However, the mechanism underlying the involvement of let-7b-5p in acute PE remains unclear. To address this, we investigated the role of let-7b-5p in acute PE in both in vitro and in vivo experimental models. The results showed that let-7b-5p upregulated the expression of stress-associated endoplasmic reticulum protein 1 (SERP1) at the post-transcriptional level. SERP1 activation leads to modulation of its chaperone protein SEC61B in the response of endoplasmic reticulum (ER) stress. Furthermore, our data show that the unfolded protein response was triggered and activation of unfolded proteins GRP78, PERK, RNF121, and CHOP occurred through the PERK-CHOP pathway, resulting in an inflammatory response and apoptosis of lung epithelial cells. These characteristics were promoted by the in vitro expression of a let-7b-5p mimic; conversely, transfection with a let-7b-5p inhibitor decreased the response of ER stress in acute PE. The results from this study thus provide evidence that let-7b-5p promotes protein processing during ER stress response by upregulating SERP1 expression, ultimately resulting in an inflammatory response and apoptosis of lung cells, cumulatively playing a critical role in the pathogenesis of acute PE.

PM2.5 triggered apoptosis in lung epithelial cells through the mitochondrial apoptotic way mediated by a ROS-DRP1-mitochondrial fission axis

Epidemiological studies revealed a sharp increase in respiratory diseases attributed to PM2.5. However, the underlying mechanisms remain unclear. Evidence suggested mitochondrion as a sensitive target upon the stimulus of PM2.5, and the centrality in the pathological processes and clinical characterization of lung diseases. To investigate cell fate and related mechanisms caused by PM2.5, we exposed human lung epithelial cells (BEAS-2B) to PM2.5 (0-100 μg/mL). Consequently, PM2.5 components were found in cytoplasm, and morphological and functional alterations in mitochondria occurred, as evidenced by loss of cristae, vacuolization and even the outer mitochondrial membrane rupture, mitochondrial membrane potential collapse, enhanced reactive oxygen species (ROS)/mtROS level, calcium overload, suppressed cellular respiration and ATP production in PM2.5-treated cells. Further, disturbed dynamics toward fission was clearly observed in PM2.5-treated mitochondria, associated with DRP1 mitochondrial translocation and phosphorylation. Besides, PM2.5 induced mitochondria-mediated apoptosis. More importantly, mechanistic results revealed ROS- and DRP1-mediated mitochondrial fission in a reciprocal way, and DRP1 inhibitor (Mdivi-1) significantly alleviated the pro-apoptotic effect of PM2.5 through reversing the activated mitochondrial apoptotic pathway. In summary, our results firstly revealed PM2.5 induced apoptosis in lung epithelial cells through a ROS-DRP1-mitochodrial fission axis-mediated mitochondrial apoptotic pathway, ultimately contributing to the onset and development of pulmonary diseases.

Amniotic fluid mesenchymal stem cells ameliorate lung alveolar epithelium regeneration and reverse lung fibrosis of chronic obstructive pulmonary disease

Pulmonary emphysema progression is attributed not only to alveolar structure loss and pulmonary regeneration impairment but also to excessive inflammatory response, proteolytic and anti‐proteolytic activity imbalance, lung epithelial cell apoptosis and abnormal lung remodeling. To ameliorate lung damage with higher efficiency through lung tissue engineering and cell therapy, predifferentiating graft cells into more restricted cell types before transplantation could enhance their ability to anatomically and functionally integrate into damaged lung. An optimal lung epithelial progenitor‐like cells (LEPLCs) predifferentiation condition has been established in eGFP‐expressing amniotic fluid mesenchymal stem cells (AFMSCs). Transplantation of LEPLCs significantly improved regeneration of lung tissue as they integrated into the lung alveolar structure, relieved airway inflammation, alleviated amount of DNA damage in alveolar cells, increased expression of growth factors such as vascular endothelial growth factor, and reduced expression of matrix metalloproteinases and lung remodeling factors when compared with the elastase‐induced pulmonary emphysema mice injected with AFMSCs. In conclusion, transplantation of predifferentiated AFMSCs through intratracheal injection showed better alveolar regeneration and reverse lung fibrosis in PPE‐induced pulmonary emphysema mice.Support or Funding InformationThis research was supported by grant MOE‐108‐S‐0023‐A from the Ministry of Education of Taiwan.

A Staphylococcus pro-apoptotic peptide induces acute exacerbation of pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic and fatal disease of unknown etiology; however, apoptosis of lung alveolar epithelial cells plays a role in disease progression. This intractable disease is associated with increased abundance of Staphylococcus and Streptococcus in the lungs, yet their roles in disease pathogenesis remain elusive. Here, we report that Staphylococcus nepalensis releases corisin, a peptide conserved in diverse staphylococci, to induce apoptosis of lung epithelial cells. The disease in mice exhibits acute exacerbation after intrapulmonary instillation of corisin or after lung infection with corisin-harboring S. nepalensis compared to untreated mice or mice infected with bacteria lacking corisin. Correspondingly, the lung corisin levels are significantly increased in human IPF patients with acute exacerbation compared to patients without disease exacerbation. Our results suggest that bacteria shedding corisin are involved in acute exacerbation of IPF, yielding insights to the molecular basis for the elevation of staphylococci in pulmonary fibrosis.

Curcumin, an Active Constituent of Turmeric Spice: Implication in the Prevention of Lung Injury Induced by Benzo(a) Pyrene (BaP) in Rats.

Benzo(a)pyrene (BaP) is a well-known carcinogen and enhances oxidative stress and apoptosis and also alters several molecular pathways. Curcumin is an active ingredient of Curcuma longa, and it has potent anti-inflammatory, antioxidant activity that defends cells from oxidative stress and cell death. The objectives of the present study were to explore the protective effects of curcumin against long-term administration of BaP induced disturbances in lungs of rats. Male rats were randomly divided into four groups: saline control, BaP only, BaP + curcumin, and curcumin only. Lung histopathology, electron microscopy, inflammatory cytokine release, antioxidant levels, apoptosis, and cell cycle were examined. Instillation of BaP significantly increased infiltration of inflammatory cells in alveolar space and inflammatory cytokine in blood. BaP induced lung tissue alterations including mild bronchitis, scant chronic inflammatory cell infiltrate in the wall of the respiratory bronchiole, and mild intra-alveolar haemorrhage. However, these alterations were found to be significantly less as mild inflammatory cell infiltrate in curcumin plus BaP treated group. Furthermore, electron microscopy results also showed necrotic changes and broken cell membrane of Type-II epithelial cell of alveoli in BaP group, which was reduced after adding curcumin treatment. In addition, we found BaP plus curcumin treatment effectively reduced inflammatory cytokines Tumour Necrosis Factor alpha (TNF-α), Interleukin 6 (IL-6), and C-reactive protein (CRP) levels in blood serum. Moreover, the levels of tunnel staining and p53 expression were significantly increased by BaP, whereas these changes were noticeably modulated after curcumin treatment. BaP also interferes in normal cell cycle, which was significantly improved with curcumin treatment. Overall, our findings suggest that curcumin attenuates BaP -induced lung injury, probably through inhibiting inflammation, oxidative stress and apoptosis in lung epithelial cells, and improving cell proliferation and antioxidants level. Thus, curcumin may be an alternative therapy for improving the outcomes of Benzo(a)pyrene-induced lung injury.

A40 Antioxidant, Anti-Inflammatory, and Antiapoptotic Potential of Curcumin in Benzo(a)pyrene (BaP)-Induced Lung Injury in Rats

Benzo(a)pyrene (BaP) is a well-known pollutant that directly induces inflammatory microenvironment in the lung. It also enhances oxidative stress and apoptosis and interferes with several other molecular pathways including cell death, survival, and proliferation that disturb normal homeostasis of the lung. Curcumin (Cur) has potent anti-inflammatory, antioxidant activity that defends cells from oxidative stress and cell death. The objectives of the present study were to explore the protective effects of curcumin against long-term administration of BaP-induced disturbances in lungs of rats. Male rats were randomly divided into four groups: saline control, BaP only, BaP + Cur, and Cur only. Lung injury histopathology, electron microscopy, inflammatory cytokine release, antioxidant levels, apoptosis, and cell cycle were examined. Instillation of BaP significantly increased infiltration of inflammatory cells in alveolar space and inflammatory cytokine in blood. Histopathologic examination found BaP-induced pulmonary inflammatory changes were improved after administration of curcumin as evident by less infiltration of macrophages and neutrophils in alveolar space, less deposition of collagen, and edema. Furthermore, electron microscopy results also showed necrotic changes and broken cell membrane of Type II epithelial cell (T2E) of alveoli in BaP group, which was reduced after addition of curcumin treatment. In addition, we found BaP plus curcumin treatment effectively reduced inflammatory cytokines TNF-α and IL-6 in blood serum, but no significant changes were found in CRP levels. Moreover, the levels of tunnel staining and p53 expression were significantly increased by BaP, whereas these changes were noticeably modulated after curcumin treatment. Bap also interferes in normal cell cycle, which was markedly improved with curcumin treatment. Overall. these findings suggest that curcumin attenuates BaP-induced lung injury, probably through inhibiting inflammation, oxidative stress, and apoptosis in lung epithelial cells, and improving cell proliferation and antioxidants' level. Thus, curcumin may be an alternative therapy for improving the outcomes of benzo(a)pyrene-induced lung injury.

Benzalkonium chloride and cetylpyridinium chloride induce apoptosis in human lung epithelial cells and alter surface activity of pulmonary surfactant monolayers

Quaternary ammonium compounds (e.g., benzalkonium chloride (BAC) and cetylpyridinium chloride (CPC)) constitute a group of cationic surfactants are widely used for personal hygiene and medical care despite the potential pulmonary toxicity. To examine whether BAC and CPC aerosols deposited in the alveolar region alter pulmonary function, we studied the effects on pulmonary surfactant using two-step in vitro models; cytotoxicity using A549 alveolar epithelial cell and changes in surface activity of the pulmonary surfactant monolayer using both Surfacten® and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Cell viability was decreased with BAC and CPC dose-dependently. A comparison of cytotoxicity among BAC homologues with different length of alkyl chain showed that C16-BAC, which has the longest alkyl chain, was more cytotoxic than C12- or C14-BAC. Caspase-3/7 activity and cleaved form of caspase-3 and PARP were increased in BAC- and CPC-exposed cells. The elevated caspase-3/7 activity and their cleaved active forms were abolished by caspase-3-inhibitor. Furthermore, we examined the features of the surface pressure/trough area (π-A) isotherm by the Langmuir-Wilhelmy method and atomic force microscopy (AFM) images of lipid monolayers on a subphase containing BAC, CPC, or pyridinium chloride (PC, as a control). The π-A isotherms showed that addition of BAC or CPC yielded dose-dependent increases in surface pressure without compression, indicating that BAC and CPC expand the isotherm to larger areas at lower pressure. The collapse pressure diminished with increasing concentration of CPC. Topographic images indicated that BAC and CPC resulted in smaller condensed lipid domains compared to the control. Conversely, PC without hydrocarbon tail group, showed no cytotoxicity and did not change the isotherms and AFM images. These results indicate that BAC and CPC cause cell death via caspase-3-dependent apoptotic pathway in A549 cells and alter the alveolar surfactant activity. These effects can be attributed to the long alkyl chain of BAC and CPC.

Overexpression of MECP2 attenuates cigarette smoke extracts induced lung epithelial cell injury by promoting CYP1B1 methylation.

MECP2 (Methyl-CpG-binding protein 2) has been shown to have a critical role in regulating DNA methylation against smoke exposed lung injury. However, the biological function of MECP2 and the underlying molecular mechanism remains elusive. Human bronchial epithelial (16HBE) and alveolar type II epithelial cells (AECII) were exposed to increasing concentrations of cigarette smoke extracts (CSE) solution to establish CSE-induced lung epithelial cell injury models. Our findings revealed that MECP2 was down-regulated, while CYP1B1 was up-regulated in CSE-induced lung epithelial cell injury models by quantitative real time PCR, western blotting and immunofluorescence staining. Down-regulated CYP1B1 was ascribed to the demethylation of its promoter by methylation-specific PCR (MSP). The in vitro experiments further showed that MECP2 overexpression significantly attenuated CSE-triggered cell growth attenuation, cell cycle arrest, apoptosis and ROS generation in lung epithelial cells by CCK-8 and flow cytometry assays. In molecular level, we further demonstrated that MECP2 overexpression obviously suppressed the expression of CYP1B1 through enhancing DNA methylation. Therefore, our data suggest that MECP2 protects against CSE-induced lung epithelial cell injury possibly through down-regulating CYP1B1 expression via elevating its methylation status.

Mesenchymal Stem Cell-Derived Exosomes and Other Extracellular Vesicles as New Remedies in the Therapy of Inflammatory Diseases.

There is growing evidence that mesenchymal stem cell (MSC)-based immunosuppression was mainly attributed to the effects of MSC-derived extracellular vesicles (MSC-EVs). MSC-EVs are enriched with MSC-sourced bioactive molecules (messenger RNA (mRNA), microRNAs (miRNAs), cytokines, chemokines, immunomodulatory factors) that regulate phenotype, function and homing of immune cells. In this review article we emphasized current knowledge regarding molecular mechanisms responsible for the therapeutic effects of MSC-EVs in attenuation of autoimmune and inflammatory diseases. We described the disease-specific cellular targets of MSC-EVs and defined MSC-sourced molecules, which were responsible for MSC-EV-based immunosuppression. Results obtained in a large number of experimental studies revealed that both local and systemic administration of MSC-EVs efficiently suppressed detrimental immune response in inflamed tissues and promoted survival and regeneration of injured parenchymal cells. MSC-EVs-based anti-inflammatory effects were relied on the delivery of immunoregulatory miRNAs and immunomodulatory proteins in inflammatory immune cells (M1 macrophages, dendritic cells (DCs), CD4+Th1 and Th17 cells), enabling their phenotypic conversion into immunosuppressive M2 macrophages, tolerogenic DCs and T regulatory cells. Additionally, through the delivery of mRNAs and miRNAs, MSC-EVs activated autophagy and/or inhibited apoptosis, necrosis and oxidative stress in injured hepatocytes, neurons, retinal cells, lung, gut and renal epithelial cells, promoting their survival and regeneration.

Regulatory mechanism of NOV/CCN3 in the inflammation and apoptosis of lung epithelial alveolar cells upon lipopolysaccharide stimulation.

Lipopolysaccharide (LPS) induces inflammatory stress and apoptosis. Pulmonary epithelial cell apoptosis has been shown to accelerate the progression of acute lung injury (ALI)/acute respiratory distress syndrome (ARDS), and is the leading cause of mortality in patients with ALI/ARDS. Nephroblastoma overexpressed (NOV; also known as CCN3), an inflammatory modulator, is reported to be a biomarker in ALI. Using an LPS-induced ALI model, we investigated the expression of CCN3 and its possible molecular mechanism involved in lung alveolar epithelial cell inflammation and apoptosis. Our data revealed that LPS treatment greatly increased the level of CCN3 in human lung alveolar type II epithelial cells (A549 cell line). The A549 cells were also transfected with a specific CCN3 small interfering RNA (siRNA). CCN3 knockdown not only largely attenuated the expression of inflammatory cytokines, interleukin (IL)-1β and transforming growth factor (TGF)-β1, but also reduced the apoptotic rate of the A549 cells and altered the expression of apoptosis-associated proteins (Bcl-2 and caspase-3). Furthermore, CCN3 knockdown greatly inhibited the activation of nuclear factor (NF)-κB p65 in the A549 cells, and TGF-β/p-Smad and NF-κB inhibitors significantly decreased the expression level of CCN3 in A549 cells. In conclusion, our data indicate that CCN3 knockdown affects the expression of downstream genes through the TGF-β/p-Smad or NF-κB pathways, leading to the inhibition of cell inflammation and apoptosis in human alveolar epithelial cells.