Short-term and Long-term Exposure of the MucilAir™ Model to Polycyclic Aromatic Hydrocarbons

Abstract

Cells grown in monocultures are widely used to model lung tissue. As a result of these culture conditions, these cells exhibit poor morphological similarity to those present in in vivo lung tissue. MucilAir™, a 3-D in vitro model comprising human basal, goblet and ciliated cells, represents a fully differentiated respiratory epithelium that can be used as an alternative and a more realistic system. The aim of our study was to compare the effects of short-term and long-term exposure to two polycyclic aromatic hydrocarbons (PAHs) — benzo[a]pyrene (B[a]P) and 3-nitrobenzanthrone (3-NBA) — using MucilAir as a model of human lung tissue. Two concentrations (0.1 μM and 1 μM) were tested at three time points (24 hours, 7 days and 28 days). Several aspects were assessed: cytotoxicity (lactate dehydrogenase (LDH) release), integrity of the cell layer (transepithelial electrical resistance (TEER)), induction of oxidative stress (reactive oxygen species production) and changes in the expression of selected genes involved in PAH metabolism ( CYP1A1 and AKR1C2) and the antioxidant response ( ALDH3A1, SOD1, SOD2, GPX1, CAT, HMOX1 and TXNRD1). The results showed that exposure to B[a]P caused a spike in LDH release at day 5. Exposure to 3-NBA caused a number of spikes in LDH release, starting at day 5, and a decrease in TEER after 11 days. CYP1A1 gene expression was upregulated after the 7-day and 28-day B[a]P exposures, as well as after the 24-hour and 7-day 3-NBA exposures. HMOX1 and SOD1 were downregulated after both 24-hour PAH treatments. HMOX1 was upregulated after a 1-week exposure to 3-NBA. There were no significant changes in the messenger RNA (mRNA) levels of AKR1C2, ALDH3A1, TXNRD1, SOD2, GPX1 or CAT. These results illustrate the potential use of this 3-D in vitro lung tissue model in studying the effects of chronic exposure to PAHs.

The Biological Effects of Complete Gasoline Engine Emissions Exposure in a 3D Human Airway Model (MucilAirTM) and in Human Bronchial Epithelial Cells (BEAS-2B)

“…The protein encoded by this gene is involved in the metabolism of many compounds, including polycyclic aromatic hydrocarbons. In our previous study, the expression of CYP1A1 in MucilAir TM was affected by exposure to benzo[a]pyrene, although the effects were detected after longer (7 and 28 days) exposure periods [25]. Therefore, the deregulation of CYP1A1 expression probably reflects the presence of PAHs in the exhaust.…”

“…For TEER measurements, an EVOM2 ohm meter (World Precision Instruments, Sarasota, FL, USA) in combination with an STX2 electrode was used [25]. This method represents a quantitative nondestructive way of measuring the integrity of tight junction dynamics in cell culture models.…”

“…However, exposure to complete emissions was not seen in any of these studies. Unlike cellular monocultures, 3D models have a greater potential for long-term exposure studies [25]. Regardless of this, the experiments with complete emissions reported so far have all been limited to short-term treatments, usually not exceeding 6 h [10,12,14,26,27].…”

A Complete In Vitro Toxicological Assessment of the Biological Effects of Cerium Oxide Nanoparticles: From Acute Toxicity to Multi-Dose Subchronic Cytotoxicity Study

“…Thus, the use of robust models such as PHAE models is highly encouraged. This three-dimensional model is a fully differentiated and functional human respiratory model that conserves respiratory epithelial properties such as metabolic activity, mucus production, and ciliary movement and has a life-span of up to one year [ 42 , 45 , 60 , 61 ]. In addition it was demonstrated through a study combining weight of evidence from proteomics, gene expression, and protein activity that this system is physiologically more suitable for repeated exposures to toxicants [ 46 ].…”

ATLA Evolves to the Next Stage: FRAME is partnering with SAGE Publishing to take ATLA forward

“…The second paper moves on to look at the application of one particular 3-D tissue model in the testing of inhalation toxicity, and discussing the results obtained with those obtained with standard 2-D cell monocultures. 3 Finally, we move into the area of education, with a survey of Italian medical and veterinary students on their knowledge and views on conscientious objection to the use of animals in their training. The differences between the two types of student are highlighted and discussed, making interesting reading indeed.…”