Lung Epithelial Carcinoma Cells Research Articles

Inhibition of dengue virus production and cytokine/chemokine expression by ribavirin and compound A

Dengue virus (DENV) infection is a worldwide public health problem with an increasing magnitude. The severity of disease in the patients with DENV infection correlates with high viral load and massive cytokine production – the condition referred to as “cytokine storm”. Thus, concurrent inhibition of DENV and cytokine production should be more effective for treatment of DENV infection. In this study, we investigated the effects of the antiviral agent – ribavirin (RV), and the anti-inflammatory compound – compound A (CpdA), individually or in combination, on DENV production and cytokine/chemokine transcription in human lung epithelial carcinoma (A549) cells infected with DENV. Initially, the cells infected with DENV serotype 2 (DENV2) was studied. The results showed that treatment of DENV-infected cells with RV could significantly reduce both DENV production and cytokine (IL-6 and TNF-α) and chemokine (IP-10 and RANTES) transcription while treatment of DENV-infected cells with CpdA could significantly reduce cytokine (IL-6 and TNF-α) and chemokine (RANTES) transcription. Combined RV and CpdA treatment of the infected cells showed greater reduction of DENV production and cytokine/chemokine transcription. Similar results of this combined treatment were observed for infection with any one of the four DENV (DENV1, 2, 3, and 4) serotypes. These results indicate that combination of the antiviral agent and the anti-inflammatory compound offers a greater efficiency in reduction of DENV and cytokine/chemokine production, providing a new therapeutic approach for DENV infection.

The Cytotoxicity of Layered Black Phosphorus.

Black phosphorus (BP), the latest addition to the family of 2D layered materials, has attracted much interest owing to potential optoelectronics, nanoelectronics, and biomedicine applications. Little is known about its toxicity, such as whether it could be as toxic as white phosphorus. In response to the possibility of BP employment into commercial products and biomedical devices, its cytotoxicity to human lung carcinoma epithelial cells (A549) was investigated. Following a 24 h exposure of the cells with different BP concentrations, cell viability assessments were conducted using water-soluble tetrazolium salt (WST-8) and methylthiazolyldiphenyltetrazolium bromide (MTT) assays. The toxicological effects were found to be dose-dependent, with BP reducing cell viabilities to 48% (WST-8) and 34% (MTT) at 50 μg mL(-1) exposure. This toxicity was observed to be generally intermediate between that of graphene oxides and exfoliated transition-metal dichalcogenides (MoS2, WS2, WSe2). The relatively low toxicity paves the way to utilization of black phosphorus.

Suppressor of cytokine signaling 1-modulated metalloproteinases and tissue inhibitor of metalloproteinase in pulmonary fibrosis.

An imbalance between metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) affects the synthesis and degradation of extracellular matrix molecules, which have an important role in the pathogenesis of pulmonary fibrosis. Lower mRNA expression levels of suppressor of cytokine signaling 1 (SOCS1) are present in fibroblasts from the lungs of pulmonary fibrosis. However, little is currently known regarding the precise role of SOCS1 has in idiopathic pulmonary fibrosis (IPF). The present study examined the expression levels of MMPs and TIMPs in A549 human epithelial lung carcinoma cells and human embryonic lung fibroblasts (HLFs) stimulated with transforming growth factor‑β1 (TGF‑β1) in conditions of deficiency and over‑expression of SOCS1, by transfection with a lentivirus. Overexpression of SOCS1 in A549 cells and HLFs significantly inhibited the mRNA expression levels of MMP‑1, MMP‑2 and MMP‑9 (P<0.05). In the A549 cells lacking SOCS1 expression, the mRNA expression levels of TIMP‑1 were significantly higher compared with the control groups (P<0.01). Overexpression of SOCS1 partially reversed these changes. The expression levels of TIMP‑1 in the HLFs with an overexpression of SOCS1 were decreased, as compared with the SOCS1‑deficient HLFs following TGF‑β1 stimulation; however, this finding was not significant (0.24±0.01 vs. 0.53±0.02, P>0.05). The expression levels of TIMP‑2 were significantly lower in the cells overexpressing SOCS1. Conversely, the mRNA expression levels of TIMP‑2 were significantly higher in the SOCS1‑deficient A549 cells, as compared with all of the other groups (P<0.05). TIMP‑4 expression levels were elevated in the A549 cells and HLFs transfected with the SOCS1‑deficient lentivirus. The expression levels of TIMP‑4 were significantly lower in the groups overexpressing SOCS1, as compared with the other groups. These results suggest that SOCS1 may act as a suppressor of pulmonary fibrosis, by reducing the expression of MMPs and TIMPs. Therefore, SOCS1 may be a target for IPF treatment.

Inhibition of Viability of the Respiratory Epithelial Cells Using Functionalized Graphene Oxide.

The viability of A549 cells, a human lung carcinoma epithelial cell line, was evaluated after exposure to graphene oxide (GO) and its derivatives (dodecylamine GO (DA-GO), reduced GO (rGO), and sodium dodecyl sulfate rGO (SDS-rGO)). A decrease in the relative amounts of C-OH bonds and an increase in the number of C-C and C-N bonds in the C 1s spectra indicated that the reduction of GO to rGO and the surface functionalization of GO has taken place. The appearance of amine stretching bands, out-of-plane C-H stretching vibrations, and S = O stretching bands in the infrared spectra indicated the formation of DA-GO, rGO, and SDS-rGO, respectively. Low concentrations (3-25 µg/mL) of GO, rGO, and SDS-rGO were found to be mildly toxic, whereas DA-GO exhibited severe dose-dependent toxicity over the same concentration range. High concen- trations (50-400 µg/mL) of GO and all its derivatives resulted in severe toxicity to the A549 cells. It is believed that surface functionality strongly affects the viability of A549 cells.

Cytotoxicity of palladium(II) complexes with some alkyl derivates of thiosalicylic acid. Crystal structure of the bis(S-butyl-thiosalicylate)palladium(II) complex, [Pd(S-bu-thiosal)2

The spectroscopically predicted structure of the obtained bis(S-butyl-thiosalicylate)palladium(II) complex, [Pd(S-bu-thiosal)2], was confirmed by an X-ray structural study. The asymmetric unit of [Pd(S-bu-thiosal)2] consists of neutral complex molecules, where the Pd(II) ion is placed in a cis-square-planar coordination environment formed by O and S atoms of two deprotonated S-butyl-thiosalicylic acid ligands. The cytotoxic effects of the S-alkyl (R=benzyl (L1), methyl (L2), ethyl (L3), propyl (L4) and butyl (L5)) derivatives of thiosalicylic acid and the corresponding palladium(II) complexes are reported here. The analysis of cancer cell viability showed that all the tested complexes are cytotoxic to human colon carcinoma cells (HCT-116 and CaCo-2) and human lung carcinoma epithelial cells (A549). The antitumor activities of the above mentioned Pd(II) complexes are higher in comparison to the corresponding ligands.

Cytotoxicity Effects of Citrate, Glucose and Cetyltrimethyl Ammonium Chloride-capped Silver Nanoparticles in Lung Epithelial Carcinoma Cells (A549)

Silver nanoparticles (AgNPs) stabilized by trisodium citrate, glucose and cetyltrimethyl ammonium chloride (CTAC) were synthesized by the reduction of silver nitrate salt in the presence of sodium borohydride (NaBH4). The UV-visible absorption spectroscopy and Transmission Electron Microscopy (TEM) were employed to investigate the growth, stability, morphology and average size of the AgNPs. The cytotoxicity effects of the silver nanomaterials were investigated using the xCELLigence real time assay on lung epithelial carcinoma cells, A549. The results show that these nanomaterials are not cytotoxic to the cells.

Liposomal β-Glucan: Preparation, Characterization and Anticancer Activities

Context: An assortment of 1,3-β-glucans have been explored for their structural and pharmacological capabilities. The immunomodulatory effects of 1,3-β-glucans have served to fight cancer. Objectives: The aim of this work was the investigation of the physiocochemical characterizations of 1,3-β-D-glucan encapsulation of liposomes and their effects on human lung carcinoma epithelial cell line A549. aterials and Methods: β-glucan was encapsulated into liposomes composed of different lipids and cholesterol [PC, PC:Chol (6:1), DDAB:DPPC:Chol (4:2:1), DCP:DPPC:Chol (4:2:1), DPPC:Chol (6:1), DSPC:Chol (6:1), DSPC]. The liposomal formulations were prepared by the hydration-dehydration method. The mean diameter of liposomes and the polydispersity index were determined by photon correlation spectroscopy (PCS) with the use of Submicron Particle Sizer. The hemocompatibility of liposome on human erythrocytes was evaluated. The MTT assay was also performed to evaluate the A549 cells viability. Results: The mean particle size of vesicles loaded with β-glucan varied from 130.20 ± 10.5 to 180.10 ± 11.3 nm. The polydispersity index ranged from 0.66 ± 0.0004 to 0.90 ± 0.08. The encapsulation efficiencies in DDAB:DPPC:Chol, DCP:DPPC:Chol and DPPC:Chol formulations were 17.20 ± 0.08%, 23.60 ± 0.20% and 18.23 ± 0.06%, respectively. The hemolysis rates did not exceed the negative control value by more than 2% and 8% at 10 and 24 h, respectively. These three formulations showed higher growth suppression of A549 cell compared to free β-glucan. This growth suppression of A549 cell by 1 μM doxorubicin was further promoted by liposomal β-glucan. Conclusion: Liposomal 1,3-β-D-glucan might potentially enhance antitumor activity.

Is the toxic potential of nanosilver dependent on its size?

BackgroundNanosilver is one of the most commonly used engineered nanomaterials (ENMs). In our study we focused on assessing the size-dependence of the toxicity of nanosilver (Ag ENMs), utilising materials of three sizes (50, 80 and 200 nm) synthesized by the same method, with the same chemical composition, charge and coating.MethodsUptake and localisation (by Transmission Electron Microscopy), cell proliferation (Relative growth activity) and cytotoxic effects (Plating efficiency), inflammatory response (induction of IL-8 and MCP-1 by Enzyme linked immune sorbent assay), DNA damage (strand breaks and oxidised DNA lesions by the Comet assay) were all assessed in human lung carcinoma epithelial cells (A549), and the mutagenic potential of ENMs (Mammalian hprt gene mutation test) was assessed in V79-4 cells as per the OECD protocol. Detailed physico-chemical characterization of the ENMs was performed in water and in biological media as a prerequisite to assessment of their impacts on cells. To study the relationship between the surface area of the ENMs and the number of ENMs with the biological response observed, Ag ENMs concentrations were recalculated from μg/cm2 to ENMs cm2/cm2 and ENMs/cm2.ResultsStudied Ag ENMs are cytotoxic and cytostatic, and induced strand breaks, DNA oxidation, inflammation and gene mutations. Results expressed in mass unit [μg/cm2] suggested that the toxicity of Ag ENMs is size dependent with 50 nm being most toxic. However, re-calculation of Ag ENMs concentrations from mass unit to surface area and number of ENMs per cm2 highlighted that 200 nm Ag ENMs, are the most toxic. Results from hprt gene mutation assay showed that Ag ENMs 200 nm are the most mutagenic irrespective of the concentration unit expressed.ConclusionWe found that the toxicity of Ag ENMs is not always size dependent. Strong cytotoxic and genotoxic effects were observed in cells exposed to Ag ENMs 50 nm, but Ag ENMs 200 nm had the most mutagenic potential. Additionally, we showed that expression of concentrations of ENMs in mass units is not representative. Number of ENMs or surface area of ENMs (per cm2) seem more precise units with which to compare the toxicity of different ENMs.Electronic supplementary materialThe online version of this article (doi:10.1186/s12989-014-0065-1) contains supplementary material, which is available to authorized users.

Abstract 514: Heat shock transcription factor-1 and heat shock protein 60 are important for lung carcinoma anoikis resistance

Abstract Metastasis of cancer cells to distant sites leads to poor prognosis and comprises approximately 90% of cancer mortalities. It is evident that metastasis requires resistance to anoikis, an apoptotic program induced by detachment from the extracellular matrix. Heat shock proteins (HSPs) are molecular chaperones whose major functions include protein folding and suppression of apoptosis, and are transcriptionally regulated by heat shock transcription factor-1 (HSF1). HSF1 has been linked to metastasis and promotes anchorage-independent proliferation of breast cancer cells. HSP60 is a chaperone found in both the mitochondria and cytosol and is linked to metastasis and regulation of tumor cell apoptosis. To investigate whether HSF1 and HSP60 are necessary for anoikis resistance, anchorage-dependent lung carcinoma (A549, H460) and normal lung epithelial (Beas-2b) cells were suspended by plating on poly-2-hydroxyethyl-methacrylate-coated culture dishes. Anoikis induction was measured by executioner caspase-7 cleavage and PARP cleavage by western blot. Protein expression was measured by western blot. Small-interfering RNAs were used to inhibit hsf1 and hsp60. We show here that upon detachment for 24-48h, lung cancer cells resisted caspase-dependent anoikis in contrast to normal lung epithelial cells, with A549 cells showing the highest level of anoikis resistance. After 24h detachment there was increased phosphorylation of HSF1 on activating serine 326 in lung cancer cells. Antisense inhibition of hsf1 and hsp60 in attached lung cancer cells had minimal effects on caspase-7 and PARP cleavage. In contrast, hsf1 and hsp60 knockdown in cancer cells that were suspended for 24-48h initiated caspase-dependent anoikis. Furthermore, knockdown of hsf1 resulted in inhibition of HSP72, HSP40, and HSP27. HSP60 was not significantly inhibited with loss of HSF1. This data suggests that HSF1 and HSP60 are necessary for lung carcinoma anoikis resistance. Our work adds to evidence suggesting HSF1 and HSP60 are promising therapeutic targets for metastasis prevention. Although more detailed mechanistic evaluation is necessary, a therapeutic approach to induce anoikis in circulating tumor cells by modulating HSF1 or HSP60 may be promising. Citation Format: Jack D. Carter, Suzanne K. Murphy. Heat shock transcription factor-1 and heat shock protein 60 are important for lung carcinoma anoikis resistance. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 514. doi:10.1158/1538-7445.AM2014-514

Abstract 4295: A 3D culture model for screening of cancer therapeutics

Abstract 3D cell culture models are currently of great interest to the bioscience community. This is most notable in the cancer biology area, where a growing body of evidence suggests that more relevant and translational observations can be made by testing compounds in a physiological relevant 3D model compared to a 2D model. In this study, we take a kinetic, live-cell imaging approach to measure the growth or shrinkage of non-adherent tumor spheroids using Ultra Low Attachment (ULA) 96-well and 384-well microtiter plates (Corning®). These round-bottomed, hydrogel coated plates facilitate the formation of spheroids by promoting self-adherence as opposed to adherence to the plate surface. In a single cell type, monoculture model, A549 lung epithelial carcinoma cells developed into single spheroid structures in each well of 96 and 384-well plates within 48-96 hours in culture. In both plate types, the resulting spheroid size, ranging from 330 µm - 600 µm, was proportional to the number of cells (1K - 5K cells per well). Post formation, spheroids remained untreated or were subjected to three test agents, staurosporine, oxaliplatin or cycloheximide. The increase or decrease in spheroid size was monitored over 3 - 10 days using the IncuCyte™ ZOOM Live Content Imaging system. Using the red fluorescent channel, spheroid growth was analyzed using fluorescent area and intensity metrics. Whereas staurosporine and oxaliplatin significantly decreased spheroid size at high concentrations, cycloheximide treatment resulted in a cytostatic response, effectively inhibiting spheroid growth in a concentration dependent manner, while not reducing total spheroid area at any tested concentration. Interestingly, in side-by-side comparisons of kinetic 2D and 3D assays, we observed a significant, 10-fold (one log) shift in the pharmacology of cycloheximide. In a 3D co-culture breast cancer model, this study also illustrates the effect of Lapatinib on SK-BR-3 cells grown in the presence of mammary derived stromal cells (fibroblasts). Together, this study describes a kinetic, live-cell approach to measuring spheroid size over time, illustrates a differential response of cycloheximide in a 3D system compared to identically treated cells grown in a 2D monolayer, and details a complex co-culture method for elucidating the effect of Lapatinib in a physiologically relevant 3D model of breast cancer. Citation Format: Kalpana Patel, Belinda O'Clair, Tim O'Callaghan, Daniel M. Appledorn, Derek Trezise. A 3D culture model for screening of cancer therapeutics. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4295. doi:10.1158/1538-7445.AM2014-4295

Synthesis and anticancer activity evaluation of 3,4-mono- and bicyclosubstituted N-(het)aryl trifluoromethyl succinimides

Novel trifluoromethylated mono- and bicyclic succinimides derived from trifluoromethylmaleic anhydride were synthesized using cyclopentadiene or 2,3-dimethylbutadiene and (het)arylamines. The biological activity of these compounds was evaluated using prediction methods and experimental studies. This series of new trifluoromethyl succinimides (3a,b and 6a-c) were tested by the National Cancer Institute (NCI, Bethesda, USA) by Program NCI-60 DTP Human Tumor Cell Line Screen at a single high dose (10−5M). Imides revealed activity on Leukemia cell lines (RPMI-8226 - myeloma cell line), Non-Small Cell Lung Cancer cell lines (A549/ATCC - lung carcinoma epithelial cells) and Renal cancer cell lines (A498 and SN12C).

Cytotoxicity of exfoliated transition-metal dichalcogenides (MoS2 , WS2 , and WSe2 ) is lower than that of graphene and its analogues.

Studies involving transition-metal dichalcogenides (TMDs) have been around for many decades and in recent years, many were focused on using TMDs to synthesize inorganic analogues of carbon nanotubes, fullerene, as well as graphene and its derivatives with the ultimate aim of employing these materials into consumer products. In view of this rising trend, we investigated the cytotoxicity of three common exfoliated TMDs (exTMDs), namely MoS2 , WS2 , and WSe2 , and compared their toxicological effects with graphene oxides and halogenated graphenes to find out whether these inorganic analogues of graphenes and derivatives would show improved biocompatibility. Based on the cell viability assessments using methylthiazolyldiphenyl-tetrazolium bromide (MTT) and water-soluble tetrazolium salt (WST-8) assays on human lung carcinoma epithelial cells (A549) following a 24 h exposure to varying concentrations of the three exTMDs, it was concluded that MoS2 and WS2 nanosheets induced very low cytotoxicity to A549 cells, even at high concentrations. On the other hand, WSe2 exhibited dose-dependent toxicological effects on A549 cells, reducing cell viability to 31.8 % at the maximum concentration of 400 μg mL(-1) ; the higher cytotoxicity displayed by WSe2 might be linked to the identity of the chalcogen. In comparison with graphene oxides and halogenated graphenes, MoS2 and WS2 were much less hazardous, whereas WSe2 showed similar degree of cytotoxicity. Future in-depth studies should be built upon this first work on the in vitro cytotoxicity of MoS2 and WS2 to ensure that they do not pose acute toxicity. Lastly, nanomaterial-induced interference control experiments revealed that exTMDs were capable of reacting with MTT assay viability markers in the absence of cells, but not with WST-8 assay. This suggests that the MTT assay is not suitable for measuring the cytotoxicity of exTMDs because inflated results will be obtained, giving false impressions that the materials are less toxic.

RIG-I enhanced interferon independent apoptosis upon Junin virus infection.

Junin virus (JUNV) is the etiological agent of Argentine hemorrhagic fever (AHF), a human disease with a high case-fatality rate. It is widely accepted that arenaviral infections, including JUNV infections, are generally non-cytopathic. In contrast, here we demonstrated apoptosis induction in human lung epithelial carcinoma (A549), human hepatocarcinoma and Vero cells upon infection with the attenuated Candid#1 strain of, JUNV as determined by phosphatidylserine (PS) translocation, Caspase 3 (CASP3) activation, Poly (ADP-ribose) polymerase (PARP) cleavage and/or chromosomal DNA fragmentation. Moreover, as determined by DNA fragmentation, we found that the pathogenic Romero strain of JUNV was less cytopathic than Candid#1 in human hepatocarcinoma and Vero, but more apoptotic in A549 and Vero E6 cells. Additionally, we found that JUNV-induced apoptosis was enhanced by RIG-I signaling. Consistent with the previously reported role of RIG-I like helicase (RLH) signaling in initiating programmed cell death, we showed that cell death or DNA fragmentation of Candid#1-infected A549 cells was decreased upon siRNA or shRNA silencing of components of RIG-I pathway in spite of increased virus production. Similarly, we observed decreased DNA fragmentation in JUNV-infected human hepatocarcinoma cells deficient for RIG-I when compared with that of RIG-I-competent cells. In addition, DNA fragmentation detected upon Candid#1 infection of type I interferon (IFN)-deficient Vero cells suggested a type I IFN-independent mechanism of apoptosis induction in response to JUNV. Our work demonstrated for the first time apoptosis induction in various cells of mammalian origin in response to JUNV infection and partial mechanism of this cell death.

The influence of Hurricanes Katrina and Rita on the inflammatory cytokine response and protein expression in A549 cells exposed to PM2.5 collected in the Baton Rouge–Port Allen industrial corridor of Southeastern Louisiana in 2005

Hurricanes Katrina and Rita hit the coast of Louisiana in 2005 and killed more than 2000 people. The two storms resulted in a significant spike in particulate matter (PM2.5) levels across the state of Louisiana. This report focuses on PM2.5 samples collected in 2005 from two monitoring sites in the neighboring cities of Baton Rouge and Port Allen, Louisiana. Inductively coupled plasma (ICP) revealed the presence of PM2.5-adsorbed representative and Fenton-active transition metals. Gas chromatography/mass spectrometry (GC-MS) analyses revealed the presence of 23 PAH compounds. Endotoxins were also detected. Metals and endotoxins were extracted with water. PAH were extracted with dichloromethane. In order to assess cytotoxicity, aqueous PM2.5 extracts were introduced to A549 Human Epithelial Lung Carcinoma Cells. Results indicated decreased cell viability in a dose-dependent manner, with an LC50 of 235 µg/ml and 250 µg/ml, respectively, for the two sites featured here. Endotoxins alone were not cytotoxic. The concentration of reactive oxygen species (ROS) and released LDH activity increased following exposure of A549 cells to aqueous PM2.5 extracts. Fluorescence microscopy revealed apoptotic and necrotic cell death mechanisms. ELISA revealed increased secretion of primary pro-inflammatory cytokines, IL-6, IL-8, and TNF-α. Global PCR gene expression revealed up-regulation of proteins associated with the cytokine storm; e.g. interleukins, chemokines, and TNF-α. Global antibody microarray was consistent with an inflammatory response, with up-regulation of cytokines involved in the down-field activation of the caspase cascade and kinase pathways. The up-regulation of metal-redox sensitive transcription factors, NF-κβ and AP-1, is consistent with a cell death mechanism initiated by Fenton-active transition metal redox catalysis.

Cytotoxicity of halogenated graphenes

Graphene and its family of derivatives possess unique and remarkable physicochemical properties which make them valuable materials for applications in many areas like electronics, energy storage and biomedicine. In response to the possibility of its large-scale manufacturing as commercial products in the future, an investigation was conducted to determine the cytotoxicity of one particular family of graphene derivatives, the halogenated graphenes, for the first time. Halogenated graphenes were prepared through thermal exfoliation of graphite oxide in gaseous chlorine, bromine or iodine atmospheres to yield chlorine- (TRGO-Cl), bromine- (TRGO-Br) and iodine-doped graphene (TRGO-I) respectively. 24 h exposure of human lung carcinoma epithelial cells (A549) to the three halogenated graphenes and subsequent cell viability assessments using methylthiazolyldiphenyl-tetrazolium bromide (MTT) and water-soluble tetrazolium salt (WST-8) assays revealed that all the halogenated graphenes examined are rather cytotoxic at the concentrations tested (3.125 μg mL(-1) to 200 μg mL(-1)) and the effects are dose-dependent, with TRGO-Cl reducing the cell viability to as low as 25.7% at the maximum concentration of 200 μg mL(-1). Their levels of cytotoxicity can be arranged in the order of TRGO-Cl > TRGO-Br > TRGO-I, and it is suggested that the amount of halogen present in the graphene material is the determining factor for the observed trend. Control experiments were carried out to test for possible nanomaterial-induced interference as a consequence of reaction between the halogenated graphenes and the viability markers (MTT/WST-8 reagent) or binding of the formazan products under cell-free conditions. The data obtained eliminate the probability of significant influence by these interferents as the change in the normalized percentage of formazan formed is relatively small and thorough washings were performed prior to the viability assessments to reduce the amount of halogenated graphenes that could eventually interact with the MTT/WST-8 assays. More studies need to be carried out in the future to complement the results obtained in this initial study in an attempt to develop a better understanding of the health hazards that the halogenated graphenes pose.

CXCL1 Regulation in Human Pulmonary Epithelial Cells by Tumor Necrosis Factor

Background/Aims: The chemokine CXCL1 has been reported to be expressed in lung airway epithelium and non-small cell lung cancer biopsy specimens. In this study, we investigated the effects of TNF-α, an abundant cytokine detected in inflammation and various cancers, on CXCL1 release by human A549 lung carcinoma epithelial cells. Methods: CXCL1 expression was determined by ELISA and RT-PCR. TNF-α signaling was examined by western blotting. Monocyte migration was assayed by a Transwell migration system. Results: TNF-α stimulated CXCL1 release and mRNA expression, and this release was inhibited by inhibitors of JNK, p38 MAPK, PI-3K/Akt and AP-1 transcription factor. TNF-α treatment was followed by JNK, p38 MAPK and PI3K/Akt activation. However, only the JNK inhibitor could reduce the CXCL1 mRNA level, suggesting that JNK is required mainly for CXCL1 mRNA synthesis, whereas p38 MAPK and PI-3K/Akt might be responsible for CXCL1 secretion. Dexamethasone (dex) and TGF-β reduced CXCL1 secretion, with dex upregulating the expression of MAP kinase phosphatase-1 and TGF-β causing smad2/3 activation and nuclear translocation. A functional analysis showed that the released CXCL1 enhanced monocyte migration and could be abolished by a CXCL1 neutralizing antibody and CXCR antagonist. Conclusion: We demonstrate that TNF-α induces CXCL1 expression through the JNK, p38 MAPK and PI-3K/Akt signaling pathways in human pulmonary epithelial cells.

Optimizing the antitumor selectivity of PVP-Hypericin re A549 cancer cells and HLF normal cells through pulsed blue light

Photodynamic therapy (PDT) is based on the preferential accumulation of photosensitizer in cancer cells with subsequent cytotoxicity mediated by singlet oxygen production after light excitation. As photosensitizers accumulate also in the surrounding non-cancer cells, the risk of damaging them by photosensitization is a limitation of PDT. Thus, minimizing the side-effects of PDT on normal cells is one of the challenging problems in medical practice. This paper studies the PDT side-effects of PVP-Hypericin (PVP: polyvinylpyrrolidone) photosensitizer excited with continuous or pulsed irradiation, on combined cell lines of human lung carcinoma epithelial cells (A549) and normal primary human lung fibroblast cells (HLF). In vitro PDTs are performed using pulsed or continuous irradiation with irradiance intensities I(*)=1.59, 6.34 and 14.27 mW/cm(2). The LED pulse lengths L are 0.127, 1.29, 13, 54.5 and 131 ms. Then fluorescence and phototoxicity of PVP-Hypericin in the A549 cancer cells are compared with those of HLF normal cells. Although, PVP-Hypericin accumulates more in A549 cancer cells, the results show that HLF cells produce dose-dependent photoreactions in the presence of photosensitizer. PVP-Hypericin induces the most optimized anticancer efficacy with moderate side-effects for I(*)=14.27 mW/cm(2) and L=131 ms.

ABC-cassette transporter 1 (ABCA1) expression in epithelial cells in Chlamydia pneumoniae infection

ATP-binding cassette transporter A1 (ABCA1) mediates reverse cholesterol transport and innate immunity response in different cell types. We have investigated the regulation of ABCA1 expression in response to intracellular Chlamydia pneumoniae infection in A549 epithelial lung carcinoma cells. C. pneumoniae infection decreased ABCA1 expression in A549 cells, and the activity of the ABCA1 promoter was decreased. The decreased promoter activity was dependent on its E-box and GnT-box elements of the promoter. Chlamydial growth was decreased in ABCA1-silenced epithelial lung carcinoma cells. These data indicate an important role for ABCA1 in intracellular bacterial infection.

Modifying excitation light dose of novel photosensitizer PVP-Hypericin for photodynamic diagnosis and therapy

Conventional photodynamic diagnosis (PDD) and therapy (PDT) makes use of photosensitizers that are excited by continuous light irradiation of specific wavelengths. In the case of PDT, the overdose of continuous excitation may lead to an expansion of necrosis in cancer cells or morbidity in healthy surroundings. The present study involves 5-h fluorescence imaging of living human lung epithelial carcinoma cells (A549) in the presence of a novel photosensitizer, PVP-Hypericin (PVP: polyvinylpyrrolidone) to optimize the excitation light doses for PDD and PDT. A number of time-lapse imaging experiments were performed using a low-power blue LED operating in either continuous or pulsed mode. The irradiances I* were 1.59, 6.34 and 14.27mW/cm2, the pulse lengths L being 0.127, 1.29, 13, 54.5, 131 and 60,000ms. Then, the relation between irradiance, various exposure times, photobleaching and phototoxicity of PVP-Hyperycin was investigated. Results showed a nonlinear relationship between the amounts of excitation dose, cell viability and toxicity. For all experimental I*, minimal phototoxicity and photobleaching was detected when cells were exposed to brief pulses of light (L⩽13ms). On the other hand, pulsed excitation with I*=14.27mW/cm2 and L=131ms induced high percentages of apoptosis comparable to the long exposures of L=60,000ms and the continuous excitation. Thus, replacement of continuous excitation by a pulsed method seems applicable for PDT.

Nanofibrous mats coated by homocharged biopolymer-layered silicate nanoparticles and their antitumor activity

Quaternized chitosan (QC)-organic rectorite (OREC) intercalated composites based nanoparticles were immobilized on cellulose acetate nanofibrous mats by layer-by-layer deposition technique via hydrogen bonds. The key design of those mats was the utilization of all negatively charged materials to reduce the quantity of nanoparticles immobilized on the fibers and then the cytotoxicity of the mats. The intercalated structure in nanoparticles was confirmed by selected area electron diffraction. The morphology and composition of NPs-assembled nanofibrous mats were studied by field emission scanning electron microscopy and X-ray photoelectron spectroscopy. The results trended to give clues as how the assembly process varied by adding OREC. MTT assay and cell culture experiments showed that NPs-assembled nanofibrous mats were commendably compatible with normal cells and could selectively kill human lung carcinoma epithelial cells. Hemolysis test indicated these mats had excellent blood compatibility. As a result, QC-OREC NPs-assembled nanofibrous mats by homocharged deposition process can be considered as a novel alternative method for cancer therapy.