A549 cell
The A549 cell line is a human adenocarcinomic alveolar basal epithelial cell line derived from the explanted lung carcinoma tissue of a 58-year-old Caucasian male, established in 1972 through in vitro cultivation techniques.[1][2] It exhibits key properties of type II alveolar epithelial cells, including the production of lamellar bodies and pulmonary surfactant, as well as ultrastructural features such as microvilli, tight junctions, and desmosomes.[3] These characteristics, combined with its adherent epithelial morphology, hypotriploid karyotype (modal chromosome number of 66), and population doubling time of approximately 22 hours, make A549 cells a widely adopted model for investigating lung epithelial biology.[2] Originally developed as part of efforts to establish continuous cell lines from solid human tumors, A549 cells have been continuously propagated for decades, retaining tumorigenic potential in athymic nude mice where they form tumors resembling alveolar cell carcinoma.[1][3] The line is keratin-positive and serves as an effective host for transfection, supporting applications in molecular biology such as gene expression profiling and high-content screening.[2] In research, A549 cells are prominently utilized to model respiratory diseases and toxicities, including nanoparticle-induced lung damage, oxidative stress, and inflammatory responses.[4] They facilitate studies on viral infections, such as Pseudomonas aeruginosa pathogenesis, antiviral drug testing, and SARS-CoV-2 infection models, due to their representation of alveolar diffusion barriers.[5][6] Additionally, their role in cancer research encompasses evaluating chemotherapeutic agents like carboplatin, investigating stem cell properties in lung adenocarcinoma, and assessing microplastic impacts on pulmonary epithelium.[7][8][9] Cultured typically in DMEM supplemented with fetal bovine serum at 37°C and 5% CO₂, A549 cells can be grown as monolayers or in three-dimensional aggregates to enhance physiological relevance.[4]Origin and Development
Isolation and Establishment
The A549 cell line was derived from explanted tumor tissue surgically removed from the lung of a 58-year-old Caucasian male diagnosed with pulmonary adenocarcinoma in 1972.[2] This tissue provided the primary source material for establishing the line as a model for human lung carcinoma.[10] The isolation was performed by D.J. Giard and colleagues at the National Cancer Institute as part of a systematic effort to develop continuous cell lines from a series of 200 solid human tumors, aiming to create resources for cancer research and in vitro cultivation studies.[11] The explantation process involved direct culturing of minced tumor fragments to promote outgrowth of viable cells, with successful establishment achieved after initial adaptation in vitro.[12] Following explantation, the A549 cells exhibited initial growth as adherent epithelial monolayers in media supplemented with fetal bovine serum, which supported their proliferation and enabled continuous subculturing without senescence.[2] This propagation marked the line's transition to a stable, immortalized culture, retaining characteristics of the original adenocarcinoma. The establishment and early characterization were detailed in the seminal publication by Giard et al. (1973) in the Journal of the National Cancer Institute, which reported on the successful derivation of A549 alongside other tumor lines from the study cohort.[11]Historical Significance
The A549 cell line was established in 1972 by D. J. Giard and colleagues through explant culture of lung carcinomatous tissue from a 58-year-old Caucasian male, marking it as one of the first continuous human lung cancer cell lines developed to model pulmonary carcinomas amid growing interest in tumor-derived models for cancer research. This initiative was part of a broader effort at the National Cancer Institute to derive stable cell lines from solid tumors, providing a renewable resource for studying adenocarcinoma characteristics in vitro. The line's epithelial morphology and ability to form tight junctions quickly positioned it as a valuable tool for investigating lung tumor biology. Deposited at the American Type Culture Collection (ATCC) as CCL-185 in 1975, the A549 line became widely accessible to researchers worldwide, facilitating its adoption in diverse studies and standardizing its use across laboratories.[2] Over the subsequent decades, its application evolved from basic tumor modeling in the 1970s—where it supported early investigations into viral propagation, including adenovirus replication in human lung cells—to more advanced roles in the 1990s, such as chemotherapeutic drug sensitivity assays that helped identify agents effective against non-small cell lung cancer. By the 2000s, A549 had integrated into high-throughput screening platforms, notably as part of the NCI-60 panel for anticancer drug discovery, enabling rapid evaluation of thousands of compounds for cytotoxic potential and contributing to the identification of novel therapeutics. In the 2010s, advancements in culture techniques led to its adaptation in three-dimensional (3D) spheroid and organoid models, better recapitulating tumor microenvironments and improving predictive accuracy for drug responses compared to traditional 2D cultures. These milestones underscore A549's enduring role in bridging foundational cell line research with modern, physiologically relevant assays, including brief applications in virology for studying respiratory pathogen-host interactions.Biological and Genetic Characteristics
Morphology and Physiology
A549 cells exhibit an adherent, epithelial-like morphology, forming confluent monolayers of polygonal or cuboidal cells when observed under phase-contrast microscopy.[13] These characteristics reflect their origin from alveolar basal epithelial tissue and enable their use as a model for lung epithelial behavior in vitro.[2] Under optimal culture conditions, A549 cells demonstrate a population doubling time of approximately 22 hours, indicating robust proliferative capacity typical of transformed epithelial lines.[2] Physiologically, A549 cells synthesize lecithin, a key component of pulmonary surfactant, with a high proportion of disaturated fatty acids, thereby mimicking the surfactant production of type II alveolar pneumocytes.[3] This synthesis occurs via the cytidine diphosphocholine pathway and supports their relevance in studies of alveolar function.[3] Upon prolonged culture or exposure to specific stimuli, A549 cells can differentiate to form structures containing multilamellar bodies, which are lamellar inclusions associated with surfactant storage and release in type II pneumocytes.[14] These bodies are observable via electron microscopy and enhance the cells' phenotypic similarity to alveolar epithelium.[3] A549 cells show positive staining for keratin intermediate filaments, such as cytokeratin 7 and 18, confirming their epithelial origin and maintenance of cytoskeletal features characteristic of lung-derived epithelial cells.[15][16]Genetic and Molecular Profile
The A549 cell line displays a hypotriploid karyotype characterized by a modal chromosome number of 66, observed in 24% of cells, with frequent counts of 64 (22%), 65, and 67 chromosomes, and an overall range spanning 59 to 145 chromosomes.[2] This aneuploidy includes several consistent marker chromosomes present in single copies across all cells, such as der(6)t(1;6)(q11;q27), ?del(6)(p23), del(11)(q21), del(2)(q11), M4, and M5, along with double copies of der(1;11)(q11;p15) and i(5p).[2] These structural abnormalities reflect the cell line's derivation from a human lung adenocarcinoma and contribute to its genetic instability, though the core profile remains stable for authentication purposes.[2] For cell line authentication, A549 cells exhibit a distinctive short tandem repeat (STR) profile as standardized by ATCC, enabling verification of identity and detection of cross-contamination.[2] The profile, determined via polymerase chain reaction amplification of multiple loci, is as follows:| Locus | Alleles |
|---|---|
| Amelogenin | X, Y |
| CSF1PO | 10, 12 |
| D13S317 | 11 |
| D16S539 | 11, 12 |
| D5S818 | 11 |
| D7S820 | 12 |
| TH01 | 9, 10 |
| TPOX | 8, 11 |
| vWA | 16, 17 |