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Focal adhesions and assessment of cytotoxicity
T G van Kooten1, C L Klein, M Wagner
1Institute of Pathology, Johannes Gutenberg University Mainz, Germany.
Abstract:
Focal adhesions are highly ordered assemblies of transmembrane receptors, extracellular matrix proteins, and a large number of cytoplasmic proteins, including structural proteins, as well as tyrosine kinases, phosphatases, and their substrates. They are now accepted as a prime component of signal transduction. Because focal adhesions also play an important role in cell morphology and migration, it can be argued that their presence is indicative of healthy cells. This has been the reason for several research groups to conclude that biomaterials sustaining focal adhesion assembly are biocompatible. In this study we demonstrate that cells under cytotoxic stress may still be able to retain their focal adhesions. Human umbilical vein endothelial cells at passage 2 were exposed to nickel and zinc ion solutions ranging from 1 to 0.01 mM for 4 and 24 h. Cells were seeded on fibronectin precoated glass slides or in tissue culture quality 96-well plates. MTT conversion with 1 and 0.5 mM nickel and zinc was strongly depressed, indicating that these concentrations are cytotoxic. Proliferative activity was also affected by these concentrations. Cells exposed to zinc typically retracted and detached from the surface, whereas cells exposed to nickel remained on the surface without signs of retraction. Nevertheless, cells exposed to nickel were impaired to reach confluency, which was determined by cadherin-5 expression. All these data indicate that nickel ions at a sufficient concentration influence cells in a cytotoxic way. Despite this apparent cytotoxicity, focal adhesion distribution as visualized by immunofluorescence staining of vinculin was not affected. With zinc the morphological changes were accompanied by apparent fusion of focal adhesions during retraction and finally dissolution. These data indicate that the mere presence of focal adhesions does not allow a reliable statement about the functional status of a cell. On the other hand, when focal adhesions are affected it is an excellent monitor of disturbed cell function.
Insights
Focal adhesions, crucial for cell health, can persist even in cells under cytotoxic stress from nickel and zinc. Their presence alone does not guarantee cell viability, but their disruption signals impaired cell function.
Area of Science:
- Cell Biology
- Biomaterials Science
- Toxicology
Background:
- Focal adhesions are critical cellular structures involved in signal transduction, cell morphology, and migration.
- Their presence has been used as an indicator of cell health and biomaterial biocompatibility.
- However, the relationship between focal adhesion integrity and cellular functional status under stress requires further investigation.
Purpose of the Study:
- To investigate whether focal adhesions remain intact in human umbilical vein endothelial cells (HUVECs) exposed to cytotoxic concentrations of nickel and zinc ions.
- To determine if focal adhesion presence is a reliable indicator of cellular health under toxic stress.
- To assess the impact of nickel and zinc on HUVEC morphology, proliferation, and focal adhesion dynamics.
Main Methods:
- HUVECs were exposed to varying concentrations of nickel and zinc ions for 4 and 24 hours.
- Cellular cytotoxicity was assessed using MTT conversion assays.
- Cell morphology, proliferation, cadherin-5 expression, and focal adhesion distribution (visualized by vinculin staining) were analyzed.
Main Results:
- Nickel and zinc ions at concentrations of 1 and 0.5 mM induced significant cytotoxicity and affected HUVEC proliferation.
- Zinc exposure led to cell retraction and detachment, while nickel exposure caused impaired confluency but cells remained attached.
- Despite cytotoxicity, focal adhesion distribution visualized by vinculin staining remained unaffected by nickel exposure, whereas zinc exposure caused focal adhesion fusion and dissolution.
- Nickel ions were confirmed to be cytotoxic, impacting cell confluency.
Conclusions:
- The mere presence of focal adhesions does not reliably indicate a healthy cellular state when cells are under cytotoxic stress.
- Nickel and zinc ions can induce cytotoxicity while focal adhesions may remain morphologically intact.
- Disruption or alteration of focal adhesions serves as a sensitive indicator of disturbed cellular function and toxicity.