Abstract:The aim of the study was to use multiple in vitro assays to assess the effects of a model irritant, sodium dodecyl sulphate (SDS) (≤10 mM (0.29 %, w/v)), on an in vitro model of the airway, MucilAir™. The use of MucilAir™ in recovery studies was also explored.A 24 h exposure increased IL-8 release at an SDS concentration ≥0.63 mM (0.018 %, w/v). Mucin secretion increased and transepithelial electrical resistance (TEER) decreased at SDS concentrations ≥1.25 mM (0.04 %, w/v). Cytotoxicity (lactate dehydrogenase … Show more
“…We also characterized the scaffold’s ability to clear from detergent residues by SDS content assessment at different stages of washing. It is known that SDS detergent residues are cytotoxic for cells ( Gilpin et al, 2014 ; Welch et al, 2021 ), with endothelial cells being particularly susceptible ( Zvarova et al, 2016 ). As ECs are crucial for vessel repopulation, we analyzed the SDS concentration at the end of the decellularization procedure and after the application of several washing steps with PBS.…”
The use of biologically derived vessels as small-diameter vascular grafts in vascular diseases is currently intensely studied. Vessel decellularization provides a biocompatible scaffold with very low immunogenicity that avoids immunosuppression after transplantation. Good scaffold preservation is important as it facilitates successful cell repopulation. In addition, mechanical characteristics have to be carefully evaluated when the graft is intended to be used as an artery due to the high pressures the vessel is subjected to. Here, we present a new and fast decellularization protocol for porcine carotid arteries, followed by investigation of the quality of obtained vessel scaffolds in terms of maintenance of important extracellular matrix components, mechanical resistance, and compatibility with human endothelial cells. Our results evidence that our decellularization protocol minimally alters both the presence of scaffold proteins and their mechanical behavior and human endothelial cells could adhere to the scaffold in vitro. We conclude that if a suitable protocol is used, a high-quality decellularized arterial scaffold of non-human origin can be promptly obtained, having a great potential to be recellularized and used as an arterial graft in transplantation medicine.
“…We also characterized the scaffold’s ability to clear from detergent residues by SDS content assessment at different stages of washing. It is known that SDS detergent residues are cytotoxic for cells ( Gilpin et al, 2014 ; Welch et al, 2021 ), with endothelial cells being particularly susceptible ( Zvarova et al, 2016 ). As ECs are crucial for vessel repopulation, we analyzed the SDS concentration at the end of the decellularization procedure and after the application of several washing steps with PBS.…”
The use of biologically derived vessels as small-diameter vascular grafts in vascular diseases is currently intensely studied. Vessel decellularization provides a biocompatible scaffold with very low immunogenicity that avoids immunosuppression after transplantation. Good scaffold preservation is important as it facilitates successful cell repopulation. In addition, mechanical characteristics have to be carefully evaluated when the graft is intended to be used as an artery due to the high pressures the vessel is subjected to. Here, we present a new and fast decellularization protocol for porcine carotid arteries, followed by investigation of the quality of obtained vessel scaffolds in terms of maintenance of important extracellular matrix components, mechanical resistance, and compatibility with human endothelial cells. Our results evidence that our decellularization protocol minimally alters both the presence of scaffold proteins and their mechanical behavior and human endothelial cells could adhere to the scaffold in vitro. We conclude that if a suitable protocol is used, a high-quality decellularized arterial scaffold of non-human origin can be promptly obtained, having a great potential to be recellularized and used as an arterial graft in transplantation medicine.
“… 17 On the other hand, the effect of surface-active agent SDS is considered to be due to its toxicity. 44 Figure 2. Comparison of permeability enhancement between L-R5 and other PEs.…”
Section: Resultsmentioning
confidence: 99%
“… 48 However, the concentration used here is toxic at 2 mM as reported in literature. 44 The increase in permeability is therefore probably due to cell death induced by this surfactant. TEER results for C 10 would suggest that it was also toxic at a concentration of 8.5 mM ( Figure 2D ).…”
The pentapeptide L-R5 has previously been shown to transiently increase the permeability of nasal epithelial cell layers
in vitro
, allowing paracellular transport of molecules of up to 4 kDa. Protein kinase C zeta (PKC ζ), a member of a family of serine/threonine kinases was shown to be involved in tight junction modulation induced by L-R5. We show here that the ability of L-R5 to modulate tight junctions is comparable to other permeability enhancers such as bilobalide, latrunculin A or C
10
. Interaction of the peptide with the target protein occurs via electrostatic interaction, with the presence of positive charges being essential for its functionality. L-R5 is myristoylated to allow quick cell entry and onset of activity. While no epithelial cytotoxicity was detected, the hydrophobic myristoyl rest was shown to cause haemolysis. Taken together, these data show that a structural optimization of L-R5 may be possible, both from a toxicological and an efficacy point of view.
“…MucilAir™ tissues in this study showed initial TEER values of 548 ± 65 Ω × cm 2 . Similar to the EpiAirway™ tissues, there were prominent variations depending on the batch ranging from 346 ± 14 to 638 ± 81 Ω × cm 2 [ 37 ]. The difference between different batches of EpiAirway™ tissues suggests differences between donors or prolonged delivery conditions.…”
Section: Discussionmentioning
confidence: 99%
“…Similar to the values in this study, IL-6 and IL-8 values in EpiAirway™ tissues were reported as 100–350 pg/mL and 14,500 ± 3000 pg/mL, respectively [ 8 ], and 91 ± 40 pg/mL IL-6 and 5300 ± 220 pg/mL IL-8 [ 36 ]. Interbatch levels of IL-6 and IL-8 secretion in MucilAir™ can vary by a factor of two to four (IL-6: 58 ± 26–205 ± 130 pg/mL and IL-8: 4.14 ± 0.61–8.19 ± 3.29 ng/mL) [ 37 ]. MucilAir™ tissues in this study secreted up to 358 pg/mL IL-6, while IL-8 were around 100 times higher.…”
The exposure to inhaled carbon nanotubes (CNT) may have adverse effects on workers upon chronic exposure. In order to assess the toxicity of inhaled nanoparticles in a physiologically relevant manner, an air–liquid interface culture of mono and cocultures of respiratory cells and assessment in reconstructed bronchial and alveolar tissues was used. The effect of CNT4003 reference particles applied in simulated lung fluid was studied in bronchial (Calu-3 cells, EpiAirway™ and MucilAir™ tissues) and alveolar (A549 +/−THP-1 and EpiAlveolar™ +/−THP-1) models. Cytotoxicity, transepithelial electrical resistance, interleukin 6 and 8 secretion, mucociliary clearance and ciliary beating frequency were used as readout parameters. With the exception of increased secretion of interleukin 6 in the EpiAlveolar™ tissues, no adverse effects of CNT4003 particles, applied at doses corresponding to the maximum estimated lifetime exposure of workers, in the bronchial and alveolar models were noted, suggesting no marked differences between the models. Since the doses for whole-life exposure were applied over a shorter time, it is not clear if the interleukin 6 increase in the EpiAlveolar™ tissues has physiological relevance.
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