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Mini Review

Applications of A549 cell line for in vitro studies – recent advances and a summary of their receptor functions

Divya M#, Smruthi R#, Archana K# and Maddaly Ravi*

#Department of Biotechnology, Vel Tech High Tech Dr. Rangarajan Dr. Sakunthala Engineering College, Avadi, Chennai – 600062, India

*Department of Human Genetics, Faculty of Biomedical Science, Sri Ramachandra Institute of Higher Education and Research, Chennai – 600116, India

Abstract

Lung cancer prevails to be the one of the common types of cancer in both men and women. Smoking is one predominant reason behind the lung cancer though the death rate varies by sex, age and race. Among several types of lung cancers, NSCLC (Non Small Cell Lung Carcinoma) serves as the most common type accounting for about 85% of all the types. A549 cell is an adenocarcinoma human alveolar basal epithelial cell line, representing the NSCLC and is widely used for in vitro experiments. Various applications include testing the biocompatibility of lipid nanoparticles, 3D bioprinting, radiosensitization, cytotoxicity and genotoxicity assays etc. Also, this cell line possesses various receptors through which their interactive actions can be studied. The presence of histamine receptors, formyl peptide receptors, protease-activated receptors, decoy receptors, estrogen and progesterone receptors in the cell line can be mobilized to study the actions of various agonists and antagonists. We present in this mini review a glimpse of the recent applications of the A549 cell line and a systematic account its receptors along with their salient points.

Keywords: A549 cell line; 3D bioprinting; radiosensitization; histamine receptor; decoy receptor; estrogen receptor

Introduction

Lung cancer is one of the leading causes of death in both men and women. Lung cancer death rate varies significantly by sex, age, race, socio economic status because of the differences in the lifestyle patterns (Lindsey Torre et al.2016). Primarily, 85-90% of all lung cancer occurs due to smoking while the remaining lung cancer cases occur in non-smokers, mainly women (Diana Maruez et al. 2007). Although, the causes of lung cancer are almost environmental there occurs a substantial individual variation in the susceptibility to respiratory carcinogens. The synergistic interaction among risk factors may lead to substantial consequences for lung cancer risk as the etiology of lung cancer is multi-factorial (Anthony Alberg and Jonathan Samet. 2003).

Non small cell lung carcinoma accounts for 85% of all types of lung cancer other than small cell lung carcinoma. NSCLC possess three types: Squamous cell carcinoma, large cell carcinoma, adenocarcinoma (Kenfield et al. 2008). Un-repaired DNA damages caused due to cigarette smoking are likely the cause of NSCLC (Xiangde Liu et al. 2005). Any defect in the ability to properly respond and repair DNA damage would be one of the basic reasons behind many forms of cancer (Wade Harper and Stephen Elledge. 2007).

A549 cells are the adenocarcinoma human alveolar basal epithelial developed by D. J. Giard in 1972 through the removal and culturing of cancerous lung tissue from a 58 year-old Caucasian male. This cell line is used to study the development of drug therapies against lung cancer (Franklin Maryland. 2016). These cells contain certain receptors such as histamine receptors (H1R and H2R), formyl protein receptor, protease activated receptor, decoy receptor etc. Recently, researchers have found out that these cells also contain estrogen receptors (ERα and ERβ) (Jian-wei et al. 2014).

Recent applications of A549 cells in vitro

Cytotoxicity

High death rates due to lung cancer have necessitated the search of novel anti-cancer compounds such are carbazole derivatives. Carbazoles are aromatic heterocyclic compounds with anti-bacterial and anti-inflammatory activity. The novel carbazole compound (Z)-4-[9-ethyl-9aH-carbazol-3-yl) amino] pent-3-en-2-one (ECAP) demonstrates a wide range of apoptotic characteristics of A549 cells that includes DNA fragmentation and PS (Phosphatidyl serine) translocation. They also induce protein expression changes in A549 cells. Also, they induce the increase in lipid peroxidation when treated with A549 lung cancer cells. The anti-cancer properties of ECAP induced p-53 mediated apoptosis by activating the oxidative stress and decrease in expression of anti-oxidant defence proteins (Refilwe Molatlhegi et al. 2015).

Genotoxicity

Hydroquinone (HQ) has been widely used in the manufacturing of rubber, paints, motor fuels, oils, reagents for photographic developers and cosmetic products. HQ is also one of the most important components in cigarette smoke. Toxicity of HQ in human and animals is limited. HQ was found to induce the genotoxicity such as DNA damage due to oxidative stress in cell models such as hematopoietic cells and lymphocytes. The recent study investigated the expression of genes in A549 cells and found out that HQ induced 55 genes. Most of the genes expressed DNA damage age responses, oxidative stress, cell cycle arrest and change in chromosome structure, which may be the potential genotoxic effects of HQ in A549 cells. The results suggested that the limited concentration of HQ is highly toxic to A549 cells when compared to other cell lines (Chen Peng et al. 2013).

Drug discovery

Several methods of drug discovery for the A549 human lung cancer cells have attained a prominent phase in the recent trends. Bauerane, a triterpenoid is found to be an effective compound for the inhibition of proliferation of A549 cells. The cells were treated with different concentrations of bauerane and 10µM was found to be the half-maximal inhibitory concentration (IC50). This compound showed a dose dependent inhibitory action against A549 cells by inducing S-phase cell cycle arrest and apoptosis which was due to the down-regulation of P13K/AKT and STAT3 signalling pathways (Qin Chen et al. 2020).

Gene expression

The mRNA expression of RNA polymerase I subunit B (POLR1B) was measured using A549 cell lines. The gene expression analysis revealed that POLR1B was highly expressed in the cells, indicating that it may play a significant role in the regulation of lung cancer cell proliferation. Also, POLR1B depletion was found to inhibit the proliferation and induce apoptosis. Thus, POLR1B has an important function as a regulator of lung cancer, thereby facilitating the development of effective therapeutic strategies (Fan Yang et al. 2019).

Lipid nanoparticles biocompatibility

3D culture of A549 cells has been used as a model to assess the uptake of nanostructured lipid carriers (NLC). The release of LDH to the cell culture medium was assessed which reflects the cell membrane rupture. The exposure of NLCs aided in the evaluation of the release of different cytokines such as IL-1β and TNF-α, IL-8 thereby supporting the responsiveness of the model to the pro-inflammatory stimulus (Joana Magalhaes et al. 2019). 

Establishment of matrix metalloproteinases

The ex vivo 3D lung models showed nodule formation and larger nodules were detected when larger de-cellularized lung matrix was used. The lung cancer growth and metastasis can be studied with the help of secreted proteins from perfusable tumour nodule.  Thus the ex vivo 3D models are better models to study lung cancer growth than the 2D cultures (Dhruv Mishra et al. 2012). 

3D bioprinting

Tumor like lung cancer model was created using 3D printed matrix of alginate-gelatin bio-ink with A549 cells. The cells were highly viable and evenly distributed after printing and suggested that temperature change during the printing process lead to only minimal damage to the cells. The total number of cells in 3D printed matrix was high, when compared to the 2D cultured cells (Xiong Wang et al. 2018).

Utilization of NSCLC for lung cancer stem cells

Lung cancer stem cells have a significant role in the development of lung cancer. The cell lines are more commonly used than lung cancer tissues for studying the lung cancer stem cells. These stem cells serve as promising targets for cancer therapy and it is crucial to improve available techniques for their identification from normal cancer cells (Yuyi Wang et al. 2018).   

Radiosensitization

The radiosensitizing capacity of gold nanoparticles (AuNPs) and suberoylanilide hydroxamic acid (SAHA) was examined in the A549 cells. The cancer cells showed decreased colony formation and also the DNA damage was greater after combinational treatments. Thus these A549 cells helped to elucidate the synergistic radiosensitizing effect of gold nanoparticles (Nora Igaz et al. 2020).

Conclusion

A549 cell line continues to be widely used for several in vitro applications. The recent advances include their utilization for obtaining cancer stem cells, 3D bioprinting and cell/tissue engineering applications. Also, the receptors present in these cells are targets of interests along with the ligands that bind to those receptors, the transcription factors involved and the signalling pathway leading to gene expression regulation.

References

Alaa Refaat, Ahmed Abd- Rabou, Asmaa Reda. TRAIL combinations: the new ‘trail’ for cancer therapy. Oncology letters (2014), 1327-1332. DOI: 10.3892/ol.2014.1922

Anna Claudia Calvielli Castelo Branco, Fabio Seiti Yamada Yoshikawa, Anna Julia Pietrobon and Maria Notomi Sato. Role of histamine in modulating the immune response and inflammation (2018). Mediators Inflamm. DOI: 10.1155/2018/9524075

Anthony Alberg J, Jonathan Samet M. Epidemiology of lung cancer. Chest journal (2003)

Chen Peng, Dionne Arthur, Faye Liu, Jongwha Lee, Qing Xia, Martin Lavin F, Jack Ng L. Genotoxicity of hydroquinone in A549 cells. Cell Biol Toxicol (2013). DOI: 10.1007/s10565-013-9247-0

Dean Edwards P, Viroj Boonyaratanakornkit. Rapid extracellular signalling by the estrogen receptor (ER): MNAR couples ER and Src to the Map kinase signalling pathway. Molecular interventions (2003) 3(1):12-5. DOI: 10.1124/mi.3.1.12

Dhruv Mishra K, Jason Sakamoto H, Michael Thrall J, Brandi Baird N, Shanda Blackmon H, Mauro Ferrari, Jonathan Kurie M, Min Kim P. Human lung cancer cells grown in an ex vivo 3D lung model produce matrix metalloproteinases not produced in 2D culture. Plos one (2012), vol 7 issue 9

Diana Marquez-Garban C, Hsiao-Wang Chen, Michael Fishbein C, Lee Goodglick, Richard Pietras J. Estrogen receptor signalling pathways in human non-small cell lung cancer. Steroids (2007) 72 135-143. DOI: 10.1016/j.steroids.2006.11.019

en.wikipedia.org/wiki/Coagulation_factor_II_receptor

en.wikipedia.org/wiki/Formyl_peptide_receptor

Fan Yang, Haitao Liu, Junjie Zhao, Xingjie Ma, Weibo Qi. POLR1B is upregulated and promotes cell proliferation in non-small cell lung cancer. Oncology letters (2019) 19: 671-680. DOI: 10.3892/ol.2019.11136

Franklin Maryland. A549- A Model for Non Small Lung Cancer. MiBioresearch (2016)

Hannah Rohe J, Ikhlas Ahmed S, Katherine Twist E, Rolf Craven J. PGRMC1 (progesterone receptor membrane component 1): A targetable protein with multiple funtions in steroid signalling, P450 activation and drug binding. Pharmacology and therapeutics (2009) 121 14-19. DOI: 10.1016/j.pharmthera.2008.09.006

Hiromichi Niikawa, Takashi Suzuki, Yasuhiro Miki, Satoshi Suzuki, Shuji Nagasaki, Junichi Akahira, Seijiro Honma, Dean Evans B, Shin-ichi Hayashi, Takashi Kondo, Hironobu Sasano. Intratumoral estrogens and estrogen receptors in human non-small cell lung carcinoma. Clin Cancer Res (2008) 14(14)

Jian-wei Huang, Bao-zhang Guan, Liang-hong Yin, Fan-na Liu, Bo Hu, Qi-yi Zheng, Fo-lan Li, Ying-xue Zhong, Yu Chen. Effects of Estrogen related receptor alpha (ERRα) on proliferation and metastasis of human lung cancer A549 cells. J Huazhong Univ Sci Technol [Med Sci] (2014) 34 (6): 875-881. DOI: 10.1007/s11596-014-1367-0

Joana Magalhaes, Marina Pinheiro, Barbara Drasier, Dedy Septiadi, Alke Petri Fink, Susana Santos G, Barbara Rothen Rutishauser, Salette Reis. Lipid nanoparticles biocompatibility and cellular uptake in a 3D human lung model. Nanomedicine (2019). DOI: 10.2217/nnm-2019-0256

Kenfield S A, Wei E K, Stampfer M J, Rosner B A, Colditz G A. Comparison of aspects of smoking among the four histological types of lung cancer. Tobacco control (2008) 17: 198-204. DOI: 10.1136/tc.2007.022582

Lindsey Torre A, Rebecca Siegel L, Ahmedin Jemal. Lung cancer statistics. Advances in Experimental Medicine and Biology (2016) 893. DOI: 10.1007/978-3-319-24223-1_1

Nora Igaz, Krisztina Szoke, David Kovacs, Andrea Buhala, Zoltan Varga, Peter Belteky, Zsolt Razga, Laszlo Tiszlavicz, Csaba Vizler, Katalin Hideghety, Zoltan Konya, Monika Kiricsa. Synergistics radiosensitization by gold nanoparticles and the histone deacetylase inhibitor SAHA in 2D and 3D cancer cell cultures. Nanomaterials (2020), 10, 158. DOI: 10.3390/nano10010158

Qin Chen, Ming Wang, Chengji Shen. Bauerane induces S-phase cell cycle arrest, apoptosis, and inhibition of proliferation of A549 human lung cancer cells through the phosphoinositide 3-kinase (P13K)/AKT and signal transducer and activator of transcription 3 (STAT3) signalling pathway. Medical Science Monitor (2020). DOI: 10.12659/MSM.919558

Refilwe Molatlhegi P, Alisa Phulukdaree, Krishnan Anand, Robert Gengan M, Charlette Tiloke, Anil Chuturgoon A. Cytotoxic effect of a novel synthesized carbazole compound on A549 lung cancer cell line. Plos one (2015). DOI: 10.1371/journal.pone.0129874

Sarah Shirley, Alexandre Morizot, Olivier Micheau. Regulating TRAIL receptor- induced cell death at the membrane: A deadly discussion. Recent patents on anti-cancer drug discovery (2011), 6, 311-323. DOI: 10.2174/157489211796957757

Ursula Rescher, Antje Danielczyk, Arseni Markoff, Volker Gerke. Functional activation of the formyl peptide receptor by a new endogenous ligand in human lung A549 cells. J.Immunol (2002), 169:1500-1504. DOI: 10.4049/jimmunol.169.3.1500

Wade Harper J, Stephen Elledge J. The DNA damage responses: Ten years after. Molecular Cell (2007) 28. DOI: 10.1016/j.molcel.2007.11.015

Xiangde Liu, Heather Conner, Tetsu Kobayashi, , Huijung Kim, Fuqiang Wen, Shinji Abe, Quihong Fang, Xingqi Wang, Mitsuyoshi Hashimoto, Peter Bitterman, Stephen Rennard I. Cigarette smoke extract induces DNA damage but not apoptosis in human bronchial epithelial cells. American Journal of Respiratory cell and Molecular biology (2005) vol 33. DOI: 10.1165/rcmb.2003-0341OC

Xiong Wang, Xinzhi Zhang, Xingliang Dai, Xuanzhi Wang, Xinda Li, Jinfu Diao, Tao Xu. Tumor-like lung cancer model based on 3D bioprinting. 3 Biotech (2018) 8:501. DOI: 10.1007/s13205-018-1519-1

Yuyi Wang, Ming Jiang, Chi Du, Yang Yu, Yanyang Liu, Mei Li, Feng Luo. Utilization of lung cancer cell lines for the study of lung cancer stem cells (Review). Oncology letters (2018) 15: 6791-6798. DOI: 10.3892/ol.2018.8265

Zhuo Wu, Yanyan Zeng, Mingkang Zhong, Bin Wang. Targeting A549 lung adenocarcinoma cell growth and invasion with protease activated receptor-1 siRNA. Molecular medicine reports (2014), 1787-1793. DOI: 10.3892/mmr.2014.2023

 

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