What Are 16HBE Cells? Uses in Respiratory Research

The 16HBE cell line is a foundational tool in respiratory science, providing a stable and reproducible model of the human airway. Originating from the lining of the lung’s airways, these cells are immortalized, meaning they can divide indefinitely in a laboratory. Their primary function is to act as a stand-in for the airway’s protective barrier. This allows scientists to study this barrier in healthy and diseased states, investigating respiratory conditions and the effects of various substances on the lungs.

Origin and Key Characteristics

The 16HBE cell line was created from normal bronchial epithelial tissue taken from a one-year-old male. To overcome the natural limit on cell division, the cells were immortalized using a plasmid containing the Simian Virus 40 (SV40) large T-antigen. This genetic modification allows the cells to bypass normal cellular aging processes, granting them the ability to be cultured over many generations.

Despite this modification, 16HBE cells retain many features of the original tissue. They grow in a characteristic “cobblestone” pattern and express proteins called cytokeratins, which are structural markers of epithelial cells. One of their most important properties is the ability to form a polarized monolayer, where the cells have a distinct top (apical) and bottom (basal) side.

This polarization allows them to form structures called tight junctions between neighboring cells. These junctions seal the space between cells, creating a physical barrier that regulates the passage of ions and molecules, a function scientists can measure as transepithelial electrical resistance (TEER).

Applications in Respiratory Research

16HBE cells are used in a wide range of respiratory research. They are prominent in the study of cystic fibrosis (CF), a genetic disorder affecting the CFTR gene. Since 16HBE cells express a functional CFTR protein, which operates as a chloride ion channel, they serve as a “healthy” control to study how the channel works and to screen for corrective drugs.

The cells are also used to model inflammatory airway diseases like asthma and chronic obstructive pulmonary disease (COPD). Researchers expose the cells to pro-inflammatory molecules to simulate disease conditions, allowing for the investigation of cellular responses, including changes in barrier function.

In toxicology, 16HBE cells provide a platform for assessing the impact of inhaled substances on the airway lining. Scientists use them to study cellular damage from air pollutants, cigarette smoke, and vape smoke. In virology, these cells help researchers understand how respiratory viruses like influenza and SARS-CoV-2 infect the airways, as they express viral entry proteins such as ACE2 and TMPRSS2.

Laboratory Culturing Techniques

Culturing 16HBE cells requires precise, sterile conditions. They are maintained in an incubator at 37°C with 5% carbon dioxide and grown in flasks with a liquid nutrient solution called a culture medium, supplemented with fetal bovine serum and other growth factors.

A technique that enhances their physiological relevance is culturing at an Air-Liquid Interface (ALI). In standard submerged culture, cells are grown on the bottom of a plastic dish, completely covered by medium. For ALI culture, cells are seeded onto a porous membrane insert.

Initially, medium is provided to both the top (apical) and bottom (basolateral) chambers. Once a monolayer forms, the apical medium is removed, exposing the top surface of the cells to air while the basolateral side remains in the nutrient medium. This setup more closely replicates the environment of the human airway. ALI culture promotes cell differentiation and polarization, leading to the formation of more robust tight junctions and a barrier that better mimics the one found in vivo.

Strengths and Limitations as a Research Model

Strengths

The 16HBE cell line has several strengths. Being of human origin, they provide a more relevant system for studying human diseases compared to animal cell lines. Their immortalized nature ensures a consistent supply of cells, which supports reproducibility across experiments and laboratories. This contrasts with primary cells isolated directly from tissue, which have a limited lifespan and exhibit variability between donors.

Limitations

Researchers must also consider the cell line’s limitations. The immortalization process using the SV40 T-antigen can alter cellular processes, as the viral protein interacts with cell cycle-regulating proteins. This could change how cells respond to damage or treatments compared to non-immortalized cells. As transformed cells, they do not replicate the complexity of the native airway, which contains multiple cell types like goblet and ciliated cells.

Another limitation is the potential for genetic drift. As cells are repeatedly grown and divided, subtle genetic changes can accumulate, leading to variations between labs or over time. Some studies also note that 16HBE cells can become unstable or overgrow during extended ALI culture, limiting their use in certain long-term studies.