An In Vitro Assay to Quantify Effects of Micro- and Nano-Plastics on Human Gene Transcription

Abstract

In today’s age, plastic waste is a major problem for our environment. The decomposition of plastic waste causes widespread contamination in all types of ecosystems worldwide. Micro-plastics in the lower micrometer size range and especially nano-plastics can become internalized by cells and thus become a threat to human health. To investigate the effects of internalized micro- and nano-plastics on human gene transcription, we used an in vitro assay to quantify CREB (cAMP response element binding protein) mediated transcription. Here we show that CREB mediated gene expression was mainly but not exclusively induced by phosphorylation. In addition, the amount of CREB affected transcription was also studied. We were also able to show that the strong CREB mediated stimulation of transcription was diminished by micro- and nano-plastics in any chosen setting. This indicates a threat to human health via the deregulation of transcription induced by internalized micro- and nano-plastics. However, this established quantifiable in vitro transcription test system could help to screen for toxic substances and non-toxic alternatives.

Uptake and Cellular Effects of Polymethylmethacrylate on Human Cell Lines

“…HEK293 cells were plated at a density of 3 × 10 4 per well in a 24-well plate with PMMA nanobeads (4.5 × 10 9 beads/mL). After 72 h, cells were transfected with either one of the following plasmids: CMV-MGFP as the positive control, CRE-MGFP as the inducible gene and TATA-MGFP linked to a non-inducible region as the negative control [16]. All three plasmids are part of a Cignal Reporter Assay Kit (Qiagen, Venlo, The Netherlands) and based on the stimulation of a CREB-mediated protein kinase A (PKA) signaling pathway.…”

“…The plasmids used for transfection were derived from a reporter assay kit used for the quantitative assessment of signal transduction pathways. Previously, it has been shown that the addition of PMMA nanobeads will result in abolished in vitro gene expression [16]. Transfection using the same plasmids enables assessment of whether CREB-mediated signaling is altered in HEK293 cells due to the presence of PMMA as well.…”

“…Furthermore, we analyzed the influence of PMMA beads on signal transduction pathways. With the addition of PMMA resulting in abolished cAMP response element binding protein (CREB)-mediated in vitro transcription, altered CREB signaling after the uptake of PMMA by human cells is expected [16]. CREB is a ubiquitously expressed transcription factor and is essential for the function of a cell.…”

An evaluation of a hepatotoxicity risk induced by the microplastic polymethyl methacrylate (PMMA) using HepG2/THP-1 co-culture model

“…According to information submitted by the industry to the US Food and Drug Administration (FDA) under the Voluntary Cosmetic Registration Program (VCRP), PMMA is widely used in bath soaps, make-up, hair dyes, and personal cleaning/care products. It has been used in a wide range of cosmetic products (Becker et al 2011 ; Pellegrino et al 2023 ). Recently, PMMA microplastics have been detected in the human liver (Horvatits et al 2022 ), cardiac tissue (Yang et al 2023 ), and blood (Leslie et al 2022 ).…”

Crafting a Scientific Framework to Mitigate Microplastic Impact on Ecosystems

“…For instance, research has revealed the presence of MPs in human blood [13] and baby placenta [14], emphasizing the urgency to investigate their potential impacts on sensitive human organs, including the brain and heart [15,16]. Furthermore, studies have examined their influence on neurological behavior [17], the immune system [18,19] and disruption in gene transcription [20]. These findings, while significant, only scratch the surface, and a deeper exploration of this subject reveals the remarkable ability of MPs to adsorb various pollutants, acting as carriers into the human body and other living organisms [21,22].…”